Novel gene classifiers and uses thereof in autoimmune diseases

ABSTRACT

Disclosed herein are methods of detecting an altered gene expression levels in a subject suspected of having an autoimmune disorder. Further described herein are methods of treating an autoimmune disorder in a subject having an exhibiting an altered gene expression level.

CROSS-REFERENCE

This application is a continuation of International Application No.PCT/US19/31203 filed May 7, 2019 which claims the benefit of U.S.Provisional Application No. 62/669,297 filed May 9, 2018, whichapplication is incorporated herein by reference in its entirety.

BACKGROUND OF THE DISCLOSURE

Skin diseases are some of the most common human illnesses and representan important global burden in healthcare. Three skin diseases are in thetop ten most prevalent diseases worldwide, and eight fall into the top50. When considered collectively, skin conditions range from being thesecond to the 11th leading causes of years lived with disability.

SUMMARY OF THE DISCLOSURE

Disclosed herein, in certain embodiments, is a method of detecting thepresence of an autoimmune disease based on molecular risk factors. Insome instances, described herein is a method of detecting the presenceof psoriasis, lupus, or atopic dermatitis based on the molecular riskfactors. In some instances, also described herein is a method ofmonitoring the progression of an autoimmune disease, e.g., psoriasis,lupus, or atopic dermatitis, based on the molecular risk factors.

Disclosed herein, in certain embodiments, is a method of detecting geneexpression levels of at least two of IL-17A, IL-17F, IL-8, CXCL5,S100A9, and DEFB4A in a subject suspected of having psoriasis,comprising: (a) isolating nucleic acids from a skin sample obtained fromthe subject, where the skin sample comprises cells from the stratumcorneum; and (b) detecting the expression levels of at least two ofIL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4A by contacting theisolated nucleic acids with a set of probes that recognizes at least twoof IL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4A, and detect bindingbetween at least two of IL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4Aand the set of probes.

Disclosed herein, in certain embodiments, is a method of detecting geneexpression levels from a first gene classifier and a second geneclassifier in a subject suspected of having psoriasis, comprising: (a)isolating nucleic acids from a skin sample obtained from the subject,wherein the skin sample comprises cells from the stratum corneum; (b)detecting the expression levels of one or more genes from the first geneclassifier: IL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4A, bycontacting the isolated nucleic acids with a set of probes thatrecognizes one or more genes from the first gene classifier, and detectsbinding between one or more genes from the first gene classifier and theset of probes; and (c) detecting the expression levels of one or moregenes from the second gene classifier: IL-17C, S100A7, IL-17RA, IL-17RC,IL-23A, IL-22, IL-26, IL-24, IL-6, CXCL1, IFN-gamma, IL-31, IL-33, TNFα,LCN2, CCL20, and TNFRSF1A, by contacting the isolated nucleic acids withan additional set of probes that recognizes one or more genes from thesecond gene classifier, and detects binding between one or more genesfrom the second gene classifier and the additional set of probes.

Disclosed herein, in certain embodiments, is a method of treating asubject with an inhibitor of TNFα, IL-17A, or IL-23, wherein the subjecthas psoriasis, the method comprising the steps of: determining whetherthe subject has an altered gene expression level by: isolating nucleicacids from a skin sample comprising cells from the stratum corneum; andperforming or having performed an expression analysis on the skin sampleby contacting the isolated nucleic acids with a set of probes thatrecognizes at least two of IL-17A, IL-17F, IL-8, CXCL5, S100A9, andDEFB4A, and detect binding between at least two of IL-17A, IL-17F, IL-8,CXCL5, S100A9, and DEFB4A and the set of probes; and if the subject hasan altered gene expression level of at least two of IL-17A, IL-17F,IL-8, CXCL5, S100A9, and DEFB4A, then administer to the subject aninhibitor of TNFα, IL-17A, or IL-23 or increase the level of thetreatment with the inhibitor, and if the subject does not have analtered gene expression level of at least two of IL-17A, IL-17F, IL-8,CXCL5, S100A9, and DEFB4A, then does not administer the inhibitor ordiscontinue the treatment with the inhibitor.

Disclosed herein, in certain embodiments, is a method of detecting geneexpression levels of at least two of IL-13, IL-31, and TSLP in a subjectsuspected of having atopic dermatitis, comprising: (a) isolating nucleicacids from a skin sample obtained from the subject, where the skinsample comprises cells from the stratum corneum; and (b) detecting theexpression levels of at least two of IL-13, IL-31, and TSLP bycontacting the isolated nucleic acids with a set of probes thatrecognizes at least two of IL-13, IL-31, and TSLP, and detect bindingbetween at least two of IL-13, IL-31, and TSLP and the set of probes.

Disclosed herein, in certain embodiments, is a method of detecting geneexpression levels from a first gene classifier and a second geneclassifier in a subject suspected of having atopic dermatitis,comprising: (a) isolating nucleic acids from a skin sample obtained fromthe subject, wherein the skin sample comprises cells from the stratumcorneum; (b) detecting the expression levels of one or more genes fromthe first gene classifier: IL-13, IL-31, and TSLP, by contacting theisolated nucleic acids with a set of probes that recognizes one or moregenes from the first gene classifier, and detects binding between one ormore genes from the first gene classifier and the set of probes; and (c)detecting the expression levels of one or more genes from the secondgene classifier: IL-13R, IL-4R, IL-17, IL-22, CXCL9, CXCL10, CXCL11,S100A7, S100A8, S100A9, CCL17, CCL18, CCL19, CCL26, CCL27, and NOS2, bycontacting the isolated nucleic acids with an additional set of probesthat recognizes one or more genes from the second gene classifier, anddetects binding between one or more genes from the second geneclassifier and the additional set of probes.

Disclosed herein, in certain embodiments, is a method of treating asubject with an antibody that specifically binds to interleukin-13(IL-13) or interleukin-13 receptor (IL-13R), wherein the subject hasatopic dermatitis, the method comprising the steps of: determiningwhether the subject has an altered gene expression level by: obtainingor having obtained isolating nucleic acids from a skin sample comprisingcells from the stratum corneum; and performing or having performed anexpression analysis on the skin sample by contacting the isolatednucleic acids with a set of probes that recognizes at least two ofIL-13, IL-31, and TSLP, and detect binding between at least two ofIL-13, IL-31, and TSLP, and the set of probes; and if the subject has analtered gene expression level of at least two of IL-13, IL-31, and TSLP,then administer to the subject an antibody that specifically binds toIL-13 or IL-13R, and if the subject does not have an altered geneexpression level of at least two of IL-13, IL-31, and TSLP, then do notadminister the antibody that specifically binds to IL-13 or IL-13R.

BRIEF DESCRIPTION OF THE DRAWINGS

Various aspects of the disclosure are set forth with particularity inthe appended claims. A better understanding of the features andadvantages of the present disclosure will be obtained by reference tothe following detailed description that sets forth illustrativeembodiments, in which the principles of the disclosure are utilized, andthe accompanying drawings of which:

FIG. 1 shows the Th17 cytokine pathway

FIG. 2 shows the activation cycle in the skin from the IL-23/17 pathway,resulting in diseases such as psoriasis and atopic dermatitis.

FIG. 3 shows the multiple inflammatory pathways elevated in chronicatopic dermatitis, including Th1, Th2, Th17, and Th22.

FIG. 4 shows the structure of the epidermis with the elapsed time forcells to progress from the dermal layer to the stratum corneum beingapproximately 28 days.

FIG. 5 shows detection of various markers of stratum corneum geneexpression in psoriasis lesional and non-lesional skin.

FIG. 6 shows a number of drugs that target the IL-17/TH-17 pathway andwhere they effect the inflammation cycle.

FIG. 7 shows targets for an atopic dermatitis assay and their place inthe activation cycle.

FIG. 8 shows the percentage of subjects achieving an 75% reduction frombaseline in the Eczema Area and Severity Index (EASI-75) or IGA score ofcleared (0) or minimal (1) (IGA 0-1) score after 16 weeks treatment with300 mg dupilumab or placebo.

FIG. 9 shows the percentage of subjects achieving EASI-75 or IGA 1/0after 125 mg/week for 12 weeks treatment with Lebrikizumab or placebo.

FIG. 10 shows the percentage of subjects achieving EASI-75 or IGA 0/1 inand unselected population and subjects selected for elevated DPP-4levels following 12 weeks treatment with Tralokinumab or placebo.

FIG. 11 shows the percentage of subjects with detected expression ofIL-13 pathway constituents or receptor using adhesive patch sampling ofstratum corneum.

FIG. 12A shows expression of CCL17 in lesion and non-lesion skinscompared to healthy normal skin.

FIG. 12B shows the normalized gene expression change of CCL17 in lesionand non-lesion skins compared to healthy normal skin.

FIG. 13A shows expression of IL-13 in lesion and non-lesion skinscompared to healthy normal skin.

FIG. 13B shows the normalized gene expression change of IL-13 in lesionand non-lesion skins compared to healthy normal skin.

FIG. 14A shows expression of IL-22 in lesion and non-lesion skinscompared to healthy normal skin.

FIG. 14B shows the normalized gene expression change of IL-22 in lesionand non-lesion skins compared to healthy normal skin.

FIG. 15A shows expression of IL-23A (p19) in lesion and non-lesion skinscompared to healthy normal skin.

FIG. 15B shows the normalized gene expression change of IL-23A (p19) inlesion and non-lesion skins compared to healthy normal skin.

FIG. 16A shows expression of IL-31 in lesion and non-lesion skinscompared to healthy normal skin.

FIG. 16B normalized gene expression change of IL-31 in lesion andnon-lesion skins compared to healthy normal skin.

FIG. 17A shows expression of IL-31RA(1) in lesion and non-lesion skinscompared to healthy normal skin.

FIG. 17B shows normalized gene expression change of IL-31RA(1) in lesionand non-lesion skins compared to healthy normal skin.

FIG. 18A shows expression of IL-31RA(2) in lesion and non-lesion skinscompared to healthy normal skin.

FIG. 18B shows normalized gene expression change of IL-31RA(2) in lesionand non-lesion skins compared to healthy normal skin.

FIG. 19 shows IL-13 and IL-4 signaling pathways.

FIG. 20A shows expression of IL-13 in lesion and non-lesion skinscompared to healthy normal skin.

FIG. 20B shows normalized gene expression change of IL-13 in lesion andnon-lesion skins compared to healthy normal skin.

FIG. 21A shows expression of IL-13RA1 in lesion and non-lesion skinscompared to healthy normal skin.

FIG. 21B shows normalized gene expression change of IL-13RA1 in lesionand non-lesion skins compared to healthy normal skin.

FIG. 22A shows expression of IL-4R in lesion and non-lesion skinscompared to healthy normal skin.

FIG. 22B shows normalized gene expression change of IL-4R in lesion andnon-lesion skins compared to healthy normal skin.

FIG. 23 shows exemplary gene expression changes in AD samples comparedto normal, in this case expression of NOS2.

FIG. 24 shows the pathogenesis of Lupus.

FIG. 25 shows cytokines with increased gene expression detected in bothuninvolved non-lesional skin and psoriatic lesions.

FIG. 26 shows cytokines with decreased gene expression in uninvolvednon-lesional skin but increased gene expression in psoriatic lesions.

FIG. 27A shows expression of IL-31RA in AD lesion skin and non-lesionskin compared to expression of IL-31RA in normal skin.

FIG. 27B shows expression of IL-31RA(2) in AD lesion skin and non-lesionskin compared to expression of IL-31RA(2) in normal skin.

FIG. 27C shows expression of CCL17 in AD lesion skin and non-lesion skincompared to expression of CCL17 in normal skin.

FIG. 27D shows expression of IL-23A in AD lesion skin and non-lesionskin compared to expression of IL-23A in normal skin.

FIG. 27E shows expression of IL-4R in AD lesion skin and non-lesion skincompared to expression of IL-4R in normal skin.

FIG. 27F shows expression of IL-22 in AD lesion skin and non-lesion skincompared to expression of IL-22 in normal skin.

FIG. 27G shows expression of IL-13 in AD lesion skin and non-lesion skincompared to expression of IL-13 in normal skin.

FIG. 27H shows expression of IL-13RA1 in AD lesion skin and non-lesionskin compared to expression of IL-13 RA1 in normal skin.

FIG. 27I shows additional expression data for IL13RA1 and IL-13 in ADnon-lesion skin samples.

FIG. 28A shows total RNA yields (pg) from lesional (PSOR) andnon-lesional (NL) skin in psoriatic patients.

FIG. 28B shows gender composition of nonlesional and psoriatic groups.

FIG. 28C shows age distribution of nonlesional and psoriatic groups.

FIG. 29A shows measurement of ACTB and DEFB4A expression in lesional(PSOR) and normal skins (NS) at different RNA input levels.

FIG. 29B shows measurement of ACTB and S100A9 expression in lesional(PSOR) and normal skins (NS) at different RNA input levels.

FIG. 29C shows measurement of ACTB and IL-17A expression in lesional(PSOR) and normal skins (NS) at different RNA input levels.

FIG. 29D shows measurement of ACTB and IL-17F expression in lesional(PSOR) and normal skins (NS) at different RNA input levels.

FIG. 29E shows measurement of ACTB and IL-17C expression in lesional(PSOR) and normal skins (NS) at different RNA input levels.

FIG. 29F shows measurement of ACTB and IL-23A expression in lesional(PSOR) and normal skins (NS) at different RNA input levels.

FIG. 30 shows a heatmap of Ct values of 13 key genes from normal (NML),and psoriatic lesional (PSOR) and non-lesional (NL) skin samplescollected with adhesive patch-based devices.

FIG. 31 shows a heatmap of Ct values of 13 genes from paired lesional(PSOR) and non-lesional (NL) skins collected from the treatment naïvepsoriatic patients, with 2 distinct subgroups of gene expressions inlesional tissues (PSOR-1 and -2).

DETAILED DESCRIPTION OF THE DISCLOSURE

Autoimmune skin disorders occur when a person's own immune systemsmistakenly attacks healthy cells. Exemplary skin disorders comprise, butare not limited to, psoriasis, lupus, and atopic dermatitis. Psoriasisis a persistent and chronic skin condition that can change the lifecycle of skin cells. Psoriasis can cause cells to build up rapidly onthe surface of the skin. The extra skin cells can form thick, silveryscales and itchy, dry, red patches that are sometimes painful.

Atopic dermatitis is a chronic disease that affects the skin. In atopicdermatitis, the skin becomes extremely itchy. Scratching leads toredness, swelling, cracking, “weeping” clear fluid, and finally,crusting and scaling. In most cases, there are periods of exacerbationsfollowed by periods of remissions. Although it is difficult to identifyexactly how many people are affected by atopic dermatitis, an estimated20% of infants and young children experience symptoms of the disease.Approximately 60% of these infants continue to have one or more symptomsof atopic dermatitis in adulthood. Thus, more than 15 million people inthe United States have symptoms of the disease. The “lesion area” is theregion of the skin affected by atopic dermatitis. Generally a lesion ischaracterized by skin dryness (xerosis), redness, blisters, scabs, orany combination. A non-lesion area is not affected by atopic dermatitisor any other skin pathology.

Lupus, also known as lupus erythematosus, is an autoimmune disease,which affects multiple organs and systems in the body. An individual'sown immune system attacks various cells causing a wide variety of signsand symptoms. With regards to the skin, there are lupus-specific skinlesions and non-specific skin lesions. In some instances, lupuscomprises a spectrum of indications comprising cutaneous lupuserythematosus (CLE) on one end and systemic lupus erythematosus (SLE)(affecting other organs and systems) on the other end. In some cases,cutaneous lupus is categorized into three main entities: chroniccutaneous lupus (CCLE), subacute cutaneous lupus (SCLE) and acutecutaneous lupus (ACLE).

Systemic lupus erythematosus (SLE), is an autoimmune disease in whichthe body's immune system mistakenly attacks healthy tissue in many partsof the body. Symptoms vary between people and may be mild to severe.Common symptoms include painful and swollen joints, fever, chest pain,hair loss, mouth ulcers, swollen lymph nodes, feeling tired, and a redrash which is most commonly on the face. Often there are periods ofillness, called flares, and periods of remission during which there arefew symptoms.

In some embodiments, disclosed herein is a method of utilizing theexpression level of genes in a gene classifier to determine the presenceof an autoimmune skin disorder (e.g., psoriasis, atopic dermatitis, orlupus). In some instances, also described herein is a method of treatinga subject determined to have an autoimmune skin disorder (e.g.,psoriasis, atopic dermatitis, or lupus), based on the expression levelof genes in a gene classifier.

Psoriasis Gene Classifiers and Methods of Use

In some embodiments, disclosed herein is a method of detecting theexpression level of a gene from a gene classifier, which is associatedwith psoriasis. In some instances, the method comprises detecting theexpression level of Interleukin 17A (IL-17A), Interleukin 17F (IL-17F),Interleukin 8 (IL-8), C-X-C Motif Chemokine Ligand 5 (CXCL5), S100Calcium Binding Protein A9 (S100A9), Defensin Beta 4A (DEFB4A), or acombination thereof. In some instances, the method comprises (a)isolating nucleic acids from a skin sample obtained from the subject,wherein the skin sample (e.g., comprising cells from the stratumcorneum); and (b) detecting the expression level of IL-17A, IL-17F,IL-8, CXCL5, S100A9, DEFB4A, or a combination thereof, by contacting theisolated nucleic acids with a set of probes that recognizes IL-17A,IL-17F, IL-8, CXCL5, S100A9, DEFB4A, or a combination thereof, anddetects binding between IL-17A, IL-17F, IL-8, CXCL5, S100A9, DEFB4A, ora combination thereof and the set of probes.

In some embodiments, the method comprises detecting the expressionlevels of two or more, three or more, four or more, or five or more ofgenes from the gene classifier: IL-17A, IL-17F, IL-8, CXCL5, S100A9, andDEFB4A. In some cases, the method comprises detecting the expressionlevels of IL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4A. In somecases, the method comprises detecting the expression levels of IL-17A,IL-17F, IL-8, CXCL5, and S100A9. In some cases, the method comprisesdetecting the expression levels of IL-17A, IL-17F, IL-8, and CXCL5. Insome cases, the method comprises detecting the expression levels ofIL-17A, IL-17F, and IL-8. In some cases, the method comprises detectingthe expression levels of IL-17A, and IL-17F.

In some instances, the expression level is an upregulated geneexpression level. In some instances, the expression level is anupregulated gene expression level, compared to a gene expression levelof an equivalent gene from a control sample. In some cases, the controlsample is a normal skin sample. In some cases, the gene expression levelof IL-17A, IL-17F, IL-8, CXCL5, S100A9, DEFB4A, or a combination thereofis upregulated. In some instances, the upregulated gene expression leveloccurs in areas of skin comprising psoriatic plaques.

In some instances, the gene expression level of IL-17A, IL-17F, IL-8,CXCL5, S100A9, or DEFB4A is increased by at least 1-fold, 2-fold,3-fold, 4-fold, 5-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold,60-fold, 70-fold, 80-fold, 90-fold, 100-fold, 110-fold, 120-fold,130-fold, 150-fold, 200-fold, 300-fold, 500-fold, or more. In somecases, the gene expression level of IL-17A, IL-17F, IL-8, CXCL5, S100A9,or DEFB4A is increased by at least 10-fold. In some cases, the geneexpression level of IL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB4A isincreased by at least 20-fold. In some cases, the gene expression levelof IL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB4A is increased by atleast 30-fold. In some cases, the gene expression level of IL-17A,IL-17F, IL-8, CXCL5, S100A9, or DEFB4A is increased by at least 40-fold.In some cases, the gene expression level of IL-17A, IL-17F, IL-8, CXCL5,S100A9, or DEFB4A is increased by at least 50-fold. In some cases, thegene expression level of IL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB4Ais increased by at least 80-fold. In some cases, the gene expressionlevel of IL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB4A is increased byat least 100-fold. In some cases, the gene expression level of IL-17A,IL-17F, IL-8, CXCL5, S100A9, or DEFB4A is increased by at least130-fold. In some cases, the gene expression level of IL-17A, IL-17F,IL-8, CXCL5, S100A9, or DEFB4A is increased by at least 150-fold. Insome cases, the gene expression level of IL-17A, IL-17F, IL-8, CXCL5,S100A9, or DEFB4A is increased by at least 200-fold. In some cases, thegene expression level of IL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB4Ais increased by at least 300-fold. In some cases, the gene expressionlevel of IL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB4A is increased byat least 500-fold. In some cases, the increased gene expression level iscompared to a gene expression level of an equivalent gene from a controlsample. In some cases, the control sample is a normal skin sample. Insome instances, the up-regulated gene expression level occurs in areasof skin comprising psoriatic plaques.

In some cases, the gene expression level of IL-17A, IL-17F, IL-8, CXCL5,S100A9, or DEFB is increased by at least 10%, 20%, 30%, 40%, 50%, 60%,70%, 80%, 90%, 100%, 200%, 300%, 400%, 500%, or more. In some cases, thegene expression level of IL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB isincreased by at least 10%. In some cases, the gene expression level ofIL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB is increased by at least20%. In some cases, the gene expression level of IL-17A, IL-17F, IL-8,CXCL5, S100A9, or DEFB is increased by at least 30%. In some cases, thegene expression level of IL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB isincreased by at least 40%. In some cases, the gene expression level ofIL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB is increased by at least50%. In some cases, the gene expression level of IL-17A, IL-17F, IL-8,CXCL5, S100A9, or DEFB is increased by at least 80%. In some cases, thegene expression level of IL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB isincreased by at least 90%. In some cases, the gene expression level ofIL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB is increased by at least100%. In some cases, the gene expression level of IL-17A, IL-17F, IL-8,CXCL5, S100A9, or DEFB is increased by at least 150%. In some cases, thegene expression level of IL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB isincreased by at least 200%. In some cases, the gene expression level ofIL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB is increased by at least300%. In some cases, the gene expression level of IL-17A, IL-17F, IL-8,CXCL5, S100A9, or DEFB is increased by at least 500%. In some cases, theincreased gene expression level is compared to a gene expression levelof an equivalent gene from a control sample. In some cases, the controlsample is a normal skin sample. In some instances, the down-regulatedgene expression level occurs in areas of skin comprising psoriaticplaques.

In some embodiments, the set of probes recognizes at least one but nomore than six genes selected from IL-17A, IL-17F, IL-8, CXCL5, S100A9,and DEFB. In some cases, the set of probes recognizes IL-17A and IL-17F.In some cases, the set of probes recognizes IL-8, CXCL5, S100A9, andDEFB4A. In some cases, the set of probes recognizes IL-17A, IL-8, andDEFB4A. In some cases, the set of probes recognizes IL-17F, CXCL5, andS100A9. In some cases, the set of probes recognizes IL-17A, IL-17F,IL-8, CXCL5, S100A9, and DEFB.

In some embodiments, the method further comprises detecting theexpression levels of Interleukin 17C (IL-17C), S100 Calcium BindingProtein A7 (S100A7), Interleukin 17 Receptor A (IL-17RA), Interleukin 17Receptor C (IL-17RC), Interleukin 23 Subunit Alpha (IL-23A), Interleukin22 (IL-22), Interleukin 26 (IL-26), Interleukin 24 (IL-24), Interleukin6 (IL-6), C-X-C Motif Chemokine Ligand 1 (CXCL1), Interferon Gamma(IFN-gamma), Interleukin 31, (IL-31), Interleukin 33 (IL-33), TumorNecrosis Factor (TNFα), Lipocalin 2 (LCN2), C-C Motif Chemokine Ligand20 (CCL20), TNF Receptor Superfamily Member 1A (TNFRSF1A) or acombination thereof. In some cases, the detecting comprises contactingthe isolated nucleic acids with an additional set of probes thatrecognizes IL-17C, S100A7, IL-17RA, IL-17RC, IL-23A, IL-22, IL-26,IL-24, IL-6, CXCL1, IFN-gamma, IL-31, IL-33, TNFα, LCN2, CCL20,TNFRSF1A, or a combination thereof, and detects binding between IL-17C,S100A7, IL-17RA, IL-17RC, IL-23A, IL-22, IL-26, IL-24, IL-6, CXCL1,IFN-gamma, IL-31, IL-33, TNFα, LCN2, CCL20, TNFRSF1A, or a combinationthereof and the additional set of probes.

In some cases, the additional set of probes recognizes one but no morethan ten genes. In some cases, the additional set of probes recognizes2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16 or 17 genes selectedfrom IL-17C, S100A7, IL-17RA, IL-17RC, IL-23A, IL-22, IL-26, IL-24,IL-6, CXCL1, IFN-gamma, IL-31, IL-33, TNFα, LCN2, CCL20, and TNFRSF1A.

In some cases, the expression level of one or more genes selected fromIL-17C, S100A7, IL-17RA, IL-17RC, IL-23A, IL-22, IL-26, IL-24, IL-6,CXCL1, IFN-gamma, IL-31, IL-33, TNFα, LCN2, CCL20, and TNFRSF1A is anelevated gene expression level. In such cases, the gene expression levelis elevated by at least 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold,20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold,100-fold, 110-fold, 120-fold, 130-fold, 150-fold, 200-fold, 300-fold,500-fold, or more. In some instances, the gene expression level iselevated by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%,200%, 300%, 400%, 500%, or more. In some instances, the expression levelis compared to a gene expression level of an equivalent gene from acontrol sample. In some instances, the control sample is a normal skinsample.

In some embodiments, a method described herein further comprisesdetecting a skin region affected with psoriasis. In some cases, alsodescribed herein include a method monitoring the skin region affectedwith psoriasis, for about 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 6months, or more.

In some instances, the method has an improved specificity, of at leastor about 70%, 75%, 80%, 85%, 90%, or more than 95% when detecting thegene expression level of IL-17A, IL-17F, IL-8, CXCL5, S100A9, DEFB, or acombination thereof. In some embodiments, the specificity is at least orabout 70%, 75%, 80%, 85%, 90%, or more than 95% when detecting the geneexpression level of IL-17C, S100A7, IL-17RA, IL-17RC, IL-23A, IL-22,IL-26, IL-24, IL-6, CXCL1, IFN-gamma, IL-31, IL-33, TNFα, LCN2, CCL20,TNFRSF1A, or a combination thereof.

In some cases, the method also has an improved sensitivity. In someembodiments, the sensitivity is at least or about 70%, 75%, 80%, 85%,90%, or more than 95% when detecting the gene expression levels ofIL-17A, IL-17F, IL-8, CXCL5, S100A9, DEFB, or a combination thereof. Insome cases, the sensitivity is at least or about 70%, 75%, 80%, 85%,90%, or more than 95% when detecting the gene expression levels ofIL-17C, S100A7, IL-17RA, IL-17RC, IL-23A, IL-22, IL-26, IL-24, IL-6,CXCL1, IFN-gamma, IL-31, IL-33, TNFα, LCN2, CCL20, TNFRSF1A, or acombination thereof.

In some embodiments, a method described herein comprises detecting geneexpression levels from a first gene classifier and a second geneclassifier in a subject in need thereof, comprising: (a) isolatingnucleic acids from a skin sample obtained from the subject, wherein theskin sample (e.g., comprising cells from the stratum corneum); (b)detecting the expression levels of one or more genes from the first geneclassifier: IL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB, by contactingthe isolated nucleic acids with a set of probes that recognizes one ormore genes from the first gene classifier, and detects binding betweenone or more genes from the first gene classifier and the set of probes;and (c) detecting the expression levels of one or more genes from thesecond gene classifier: IL-17C, S100A7, IL-17RA, IL-17RC, IL-23A, IL-22,IL-6, IL-24, IL-6, CXCL1, IFN-gamma, IL-31, IL-33, TNFα, LCN2, CCL20,and TNFRSF1A, by contacting the isolated nucleic acids with anadditional set of probes that recognizes one or more genes from thesecond gene classifier, and detects binding between one or more genesfrom the second gene classifier and the additional set of probes.

In some embodiments, also provided herein is a method of treatingpsoriasis, which comprises administering one or more inhibitors.Inhibitors for inclusion in methods for treatment of psoriasis describedherein include, but are not limited to, inhibitors of TNFα, IL-17A, andIL-23. Exemplary inhibitors of TNFα include, but are not limited to,adalimumab, certolizumab, etanercept, golimumab, and infliximab.Exemplary inhibitors of IL-17A include, but are not limited to,ixekizumab (LY2439821), brodalumab (AMG 827), and secukinumab. Exemplaryinhibitors of IL-23 include, but are not limited to, guselkumab,tildrakizumab, and risankizumab.

In some cases, the inhibitor for inclusion in the methods describedherein for treatment of psoriasis is an inhibitor of TNFα. In somecases, the subject is treated with an inhibitor of TNFα such asadalimumab, certolizumab, etanercept, golimumab, or infliximab.

In some cases, the inhibitor for inclusion in the methods describedherein for treatment of psoriasis is an inhibitor of IL-17A. In somecases, the subject is treated with an inhibitor of IL-17A such asixekizumab (LY2439821), brodalumab (AMG 827), or secukinumab.

In some cases, the inhibitor for inclusion in the methods describedherein for treatment of psoriasis is an inhibitor of IL-23. In somecases, the subject is treated with an inhibitor of IL-23 such asguselkumab, tildrakizumab, or risankizumab.

Atopic Dermatitis Gene Classifiers and Methods of Use

In some embodiments, disclosed herein is a method of detecting theexpression level of a gene from a gene classifier, which is associatedwith atopic dermatitis. In some instances, the method comprisesdetecting the expression level of Interleukin 13 (IL-13), Interleukin 31(IL-31), Thymic Stromal Lymphopoietin (TSLP), or a combination thereof.In some instances, the method comprises (a) isolating nucleic acids froma skin sample obtained from the subject, wherein the skin sample (e.g.,comprising cells from the stratum corneum); and (b) detecting theexpression level of IL-13, IL-31, TSLP, or a combination thereof, bycontacting the isolated nucleic acids with a set of probes thatrecognizes IL-13, IL-31, TSLP, or a combination thereof, and detectsbinding between IL-13, IL-31, TSLP, or a combination thereof and the setof probes.

In some embodiments, the method comprises detecting the expressionlevels of two or more, or three or more of genes from the geneclassifier: IL-13, IL-31, and TSLP. In some cases, the method comprisesdetecting the expression levels of IL-13, IL-31, and TSLP. In somecases, the method comprises detecting the expression levels of IL-31 andTSLP. In some cases, the method comprises detecting the expressionlevels of IL-13 and IL-31. In some cases, the method comprises detectingthe expression levels of IL-13 and TSLP.

In some instances, the expression level is an upregulated geneexpression level. In some instances, the expression level is anup-regulated gene expression level, compared to a gene expression levelof an equivalent gene from a control sample. In some cases, the controlsample is a normal skin sample. In some cases, the gene expression levelof IL-13, IL-31, TSLP, or a combination thereof is up-regulated. In someinstances, the up-regulated gene expression level occurs in areas ofskin comprising atopic dermatitis.

In some instances, the gene expression level of IL-13, IL-31, or TSLP isincreased by at least 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold,20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold,100-fold, 110-fold, 120-fold, 130-fold, 150-fold, 200-fold, 300-fold,500-fold, or more. In some cases, the gene expression level of IL-13,IL-31, or TSLP is increased by at least 10-fold. In some cases, the geneexpression level of IL-13, IL-31, or TSLP is increased by at least20-fold. In some cases, the gene expression level of IL-13, IL-31, orTSLP is increased by at least 30-fold. In some cases, the geneexpression level of IL-13, IL-31, or TSLP is increased by at least40-fold. In some cases, the gene expression level of IL-13, IL-31, orTSLP is increased by at least 50-fold. In some cases, the geneexpression level of IL-13, IL-31, or TSLP is increased by at least80-fold. In some cases, the gene expression level of IL-13, IL-31, orTSLP is increased by at least 100-fold. In some cases, the geneexpression level of IL-13, IL-31, or TSLP is increased by at least130-fold. In some cases, the gene expression level of IL-13, IL-31, orTSLP is increased by at least 150-fold. In some cases, the geneexpression level of IL-13, IL-31, or TSLP is increased by at least200-fold. In some cases, the gene expression level of IL-13, IL-31, orTSLP is increased by at least 300-fold. In some cases, the geneexpression level of IL-13, IL-31, or TSLP is increased by at least500-fold. In some cases, the decreased gene expression level is comparedto a gene expression level of an equivalent gene from a control sample.In some cases, the control sample is a normal skin sample. In someinstances, the down-regulated gene expression level occurs in areas ofskin comprising atopic dermatitis.

In some cases, the gene expression level of IL-13, IL-31, or TSLP isincreased by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%,200%, 300%, 400%, 500%, or more. In some cases, the gene expressionlevel of IL-13, IL-31, or TSLP is increased by at least 10%. In somecases, the gene expression level of IL-13, IL-31, or TSLP is increasedby at least 20%. In some cases, the gene expression level of IL-13,IL-31, or TSLP is increased by at least 30%. In some cases, the geneexpression level of IL-13, IL-31, or TSLP is increased by at least 40%.In some cases, the gene expression level of IL-13, IL-31, or TSLP isincreased by at least 50%. In some cases, the gene expression level ofIL-13, IL-31, or TSLP is increased by at least 80%. In some cases, thegene expression level of IL-13, IL-31, or TSLP is increased by at least90%. In some cases, the gene expression level of IL-13, IL-31, or TSLPis increased by at least 100%. In some cases, the gene expression levelof IL-13, IL-31, or TSLP is increased by at least 150%. In some cases,the gene expression level of IL-13, IL-31, or TSLP is increased by atleast 200%. In some cases, the gene expression level of IL-13, IL-31, orTSLP is increased by at least 300%. In some cases, the gene expressionlevel of IL-13, IL-31, or TSLP is increased by at least 500%. In somecases, the decreased gene expression level is compared to a geneexpression level of an equivalent gene from a control sample. In somecases, the control sample is a normal skin sample. In some instances,the down-regulated gene expression level occurs in areas of skincomprising atopic dermatitis.

In some embodiments, the set of probes recognizes at least one but nomore than three genes selected from IL-13, IL-31, and TSLP. In somecases, the set of probes recognizes IL-13 and IL-31. In some cases, theset of probes recognizes IL-31 and TSLP. In some cases, the set ofprobes recognizes IL-13 and TSLP. In some cases, the set of probesrecognizes IL-13, IL-31, and TSLP.

In some embodiments, the method further comprises detecting theexpression levels of Interleukin 13 Receptor (IL-13R), Interleukin 4Receptor (IL-4R), Interleukin 17 (IL-17), Interleukin 22 (IL-22), C-X-CMotif Chemokine Ligand 9 (CXCL9), C-X-C Motif Chemokine Ligand 10(CXCL10), C-X-C Motif Chemokine Ligand 10 (CXCL11), S100 Calcium BindingProtein A7 (S100A7), S100 Calcium Binding Protein A8 (S100A8), S100Calcium Binding Protein A9 (S100A9), C-C Motif Chemokine Ligand 17(CCL17), C-C Motif Chemokine Ligand 18 (CCL18), C-C Motif ChemokineLigand 19 (CCL19), C-C Motif Chemokine Ligand 26 (CCL26), C-C MotifChemokine Ligand 27 (CCL27), Nitric Oxide Synthetase 2 (NOS2) or acombination thereof. In some cases, the detecting comprises contactingthe isolated nucleic acids with an additional set of probes thatrecognizes IL-13R, IL-4R, IL-17, IL-22, CXCL9, CXCL10, CXCL11, S100A7,S100A8, S100A9, CCL17, CCL18, CCL19, CCL26, CCL27, NOS2, or acombination thereof, and detects binding between IL-13R, IL-4R, IL-17,IL-22, CXCL9, CXCL10, CXCL11, S100A7, S100A8, S100A9, CCL17, CCL18,CCL19, CCL26, CCL27, NOS2, or a combination thereof and the additionalset of probes.

In some cases, the additional set of probes recognizes one but no morethan ten genes. In some cases, the additional set of probes recognizes2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16 genes selectedfrom IL-13R, IL-4R, IL-17, IL-22, CXCL9, CXCL10, CXCL11, S100A7, S100A8,S100A9, CCL17, CCL18, CCL19, CCL26, CCL27, and NOS2.

In some cases, the expression level of one or more genes selected fromIL-13R, IL-4R, IL-17, IL-22, CXCL9, CXCL10, CXCL11, S100A7, S100A8,S100A9, CCL17, CCL18, CCL19, CCL26, CCL27, and NOS2 is an elevated geneexpression level. In such cases, the gene expression level is elevatedby at least 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 20-fold,30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold,110-fold, 120-fold, 130-fold, 150-fold, 200-fold, 300-fold, 500-fold, ormore. In some instances, the gene expression level is elevated by atleast 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, 300%,400%, 500%, or more. In some instances, the expression level is comparedto a gene expression level of an equivalent gene from a control sample.In some instances, the control sample is a normal skin sample.

In some embodiments, a method described herein further comprisesdetecting a skin region affected with atopic dermatitis. In some cases,also described herein include a method monitoring the skin regionaffected with atopic dermatitis, for about 1 week, 2 weeks, 3 weeks, 1month, 2 months, 6 months, or more.

In some instances, the method has an improved specificity, of at leastor about 70%, 75%, 80%, 85%, 90%, or more than 95% when detecting thegene expression level of IL-13, IL-31, TSLP, or a combination thereof.In some embodiments, the specificity is at least or about 70%, 75%, 80%,85%, 90%, or more than 95% when detecting the gene expression level ofIL-13R, IL-4R, IL-17, IL-22, CXCL9, CXCL10, CXCL11, S100A7, S100A8,S100A9, CCL17, CCL18, CCL19, CCL26, CCL27, NOS2, or a combinationthereof.

In some cases, the method also has an improved sensitivity. In someembodiments, the sensitivity is at least or about 70%, 75%, 80%, 85%,90%, or more than 95% when detecting the gene expression levels ofIL-13, IL-31, TSLP, or a combination thereof. In some cases, thesensitivity is at least or about 70%, 75%, 80%, 85%, 90%, or more than95% when detecting the gene expression levels of IL-13R, IL-4R, IL-17,IL-22, CXCL9, CXCL10, CXCL11, S100A7, S100A8, S100A9, CCL17, CCL18,CCL19, CCL26, CCL27, NOS2, or a combination thereof.

In some embodiments, a method described herein comprises detecting geneexpression levels from a first gene classifier and a second geneclassifier in a subject in need thereof, comprising: (a) isolatingnucleic acids from a skin sample obtained from the subject, wherein theskin sample (e.g., comprising cells from the stratum corneum); (b)detecting the expression levels of one or more genes from the first geneclassifier: IL-13, IL-31, and TSLP, by contacting the isolated nucleicacids with a set of probes that recognizes one or more genes from thefirst gene classifier, and detects binding between one or more genesfrom the first gene classifier and the set of probes; and (c) detectingthe expression levels of one or more genes from the second geneclassifier: IL-13R, IL-4R, IL-17, IL-22, CXCL9, CXCL10, CXCL11, S100A7,S100A8, S100A9, CCL17, CCL18, CCL19, CCL26, CCL27, and NOS2, bycontacting the isolated nucleic acids with an additional set of probesthat recognizes one or more genes from the second gene classifier, anddetects binding between one or more genes from the second geneclassifier and the additional set of probes.

Provided herein are methods for treatment of atopic dermatitiscomprising administering one or more inhibitors described herein.Inhibitors for inclusion in methods for treatment of atopic dermatitisdescribed herein include, but are not limited to, inhibitors of IL-13,PDE4, or IL-31. In some cases, the inhibitor for inclusion in themethods described herein for treatment of atopic dermatitis is aninhibitor of IL-13. In some cases, the inhibitor for inclusion in themethods described herein for treatment of atopic dermatitis is aninhibitor of PDE4. In some cases, the inhibitor for inclusion in themethods described herein for treatment of atopic dermatitis is aninhibitor of IL-31.

In some cases, the inhibitor of IL-13 includes, but is not limited to,lebrikizumab and tralokinumab. In some cases, the inhibitor of IL-13 islebrikizumab. In some cases, the inhibitor of IL-13 is tralokinumab. Insome cases, a subject is treated with an inhibitor of IL-13 such aslebrikizumab or tralokinumab.

In some instances, the PDE4 inhibitor includes, but is not limited to,crisaborole. In some instances, a subject is treated with a PDE4inhibitor such as crisaborole.

In some instances, the IL-31 inhibitor includes, but is not limited to,nemolizumab. In some instances, a subject is treated with an IL-31inhibitor such as nemolizumab.

Lupus Gene Classifiers and Methods of Use

In some embodiments, disclosed herein is a method of detecting theexpression level of a gene from a gene classifier, which is associatedwith lupus erythematosus. In some instances, the method comprisesdetecting the expression level of Interferon Alpha 1 (IFNA1), InterferonAlpha 2 (IFNA2), Interferon Alpha 4 (IFNA4), Interferon Alpha And BetaReceptor Subunit 1 (IFNR1), Interferon Alpha And Beta Receptor Subunit 2(IFNR2), C-C Motif Chemokine Ligand 5 (CCL5), or a combination thereof.In some instances, the method comprises (a) isolating nucleic acids froma skin sample obtained from the subject, wherein the skin sample (e.g.,comprising cells from the stratum corneum); and (b) detecting theexpression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, CCL5, or acombination thereof, by contacting the isolated nucleic acids with a setof probes that recognizes IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, CCL5, or acombination thereof, and detects binding between IFNA1, IFNA2, IFNA4,IFNR1, IFNR2, CCL5, or a combination thereof and the set of probes.

In some embodiments, the method comprises detecting the expressionlevels of two or more, three or more, four or more, or five or more ofgenes from the gene classifier: IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, andCCL5. In some cases, the method comprises detecting the expressionlevels of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, and CCL5. In some cases,the method comprises detecting the expression levels of IFNA1, IFNA2,IFNA4, IFNR1, and IFNR2. In some cases, the method comprises detectingthe expression levels of IFNA1, IFNA2, IFNA4, and IFNR1. In some cases,the method comprises detecting the expression levels of IFNA1, IFNA2,and IFNA4. In some cases, the method comprises detecting the expressionlevels of IFNA1, IFNA4, and CCL5.

In some instances, the expression level is an upregulated geneexpression level. In some instances, the expression level is anup-regulated gene expression level, compared to a gene expression levelof an equivalent gene from a control sample. In some cases, the controlsample is a normal skin sample. In some cases, the gene expression levelof IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, CCL5, or a combination thereof isup-regulated. In some instances, the up-regulated gene expression leveloccurs in areas of skin comprising lupus lesions.

In some instances, the gene expression level of IFNA1, IFNA2, IFNA4,IFNR1, IFNR2, or CCL5 is increased by at least 1-fold, 2-fold, 3-fold,4-fold, 5-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 60-fold,70-fold, 80-fold, 90-fold, 100-fold, 110-fold, 120-fold, 130-fold,150-fold, 200-fold, 300-fold, 500-fold, or more. In some cases, the geneexpression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 isincreased by at least 10-fold. In some cases, the gene expression levelof IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 is increased by at least20-fold. In some cases, the gene expression level of IFNA1, IFNA2,IFNA4, IFNR1, IFNR2, or CCL5 is increased by at least 30-fold. In somecases, the gene expression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2,or CCL5 is increased by at least 40-fold. In some cases, the geneexpression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 isincreased by at least 50-fold. In some cases, the gene expression levelof IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 is increased by at least80-fold. In some cases, the gene expression level of IFNA1, IFNA2,IFNA4, IFNR1, IFNR2, or CCL5 is increased by at least 100-fold. In somecases, the gene expression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2,or CCL5 is increased by at least 130-fold. In some cases, the geneexpression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 isincreased by at least 150-fold. In some cases, the gene expression levelof IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 is increased by at least200-fold. In some cases, the gene expression level of IFNA1, IFNA2,IFNA4, IFNR1, IFNR2, or CCL5 is increased by at least 300-fold. In somecases, the gene expression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2,or CCL5 is increased by at least 500-fold. In some cases, the increasedgene expression level is compared to a gene expression level of anequivalent gene from a control sample. In some cases, the control sampleis a normal skin sample. In some instances, the up-regulated geneexpression level occurs in areas of skin comprising lupus lesions.

In some cases, the gene expression level of IFNA1, IFNA2, IFNA4, IFNR1,IFNR2, or CCL5 is increased by at least 10%, 20%, 30%, 40%, 50%, 60%,70%, 80%, 90%, 100%, 200%, 300%, 400%, 500%, or more. In some cases, thegene expression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 isincreased by at least 10%. In some cases, the gene expression level ofIFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 is increased by at least 20%.In some cases, the gene expression level of IFNA1, IFNA2, IFNA4, IFNR1,IFNR2, or CCL5 is increased by at least 30%. In some cases, the geneexpression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 isincreased by at least 40%. In some cases, the gene expression level ofIFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 is increased by at least 50%.In some cases, the gene expression level of IFNA1, IFNA2, IFNA4, IFNR1,IFNR2, or CCL5 is increased by at least 80%. In some cases, the geneexpression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 isincreased by at least 90%. In some cases, the gene expression level ofIFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 is increased by at least100%. In some cases, the gene expression level of IFNA1, IFNA2, IFNA4,IFNR1, IFNR2, or CCL5 is increased by at least 150%. In some cases, thegene expression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 isincreased by at least 200%. In some cases, the gene expression level ofIFNA1, IFNA2, IFNA4, IFNR1, IFNR2, or CCL5 is increased by at least300%. In some cases, the gene expression level of IFNA1, IFNA2, IFNA4,IFNR1, IFNR2, or CCL5 is increased by at least 500%. In some cases, thedecreased gene expression level is compared to a gene expression levelof an equivalent gene from a control sample. In some cases, the controlsample is a normal skin sample. In some instances, the up-regulated geneexpression level occurs in areas of skin comprising atopic dermatitis.

In some embodiments, the set of probes recognizes at least one but nomore than six genes selected from IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, andCCL5. In some cases, the set of probes recognizes IFNA1, IFNA2, andIFNA4. In some cases, the set of probes recognizes IFNR1, IFNR2, andCCL5. In some cases, the set of probes recognizes IFNA1, IFNA4, andIFNR2. In some cases, the set of probes recognizes IFNA2, IFNR1, andCCL5.

In some embodiments, the method further comprises detecting theexpression levels of Interferon Beta 1 (IFNB1), Interferon Epsilon(IFNE), Interferon Omega 1 (IFNW1), Adenosine Deaminase, RNA Specific(ADAR), Interferon Induced proteins with Tetratricopeptide repeat(IFIT), interferon-inducible p200 family of proteins (IFI), InterferonRegulatory Factors (IRF), 2′-5′-Oligoadenylate Synthetase 1 (OAS1),Interleukin 1 Receptor Associated Kinase 1 (IRAK1), TNF Alpha InducedProtein 3 (TNFAIP3), Autophagy Related 5 (ATG5), Tyrosine Kinase 2(TYK2), Signal Transducer and Activator Of Transcription 4 (STAT4),Osteopontin (OPN), Keratins (KRT), or a combination thereof. In somecases, the detecting comprises contacting the isolated nucleic acidswith an additional set of probes that recognizes IFNB1, IFNE, IFNW1,ADAR, IFIT, IFI, IRF, OAS1, IRAK1, TNFAIP3, ATG5, TYK2, STAT4, OPN, KRT,or a combination thereof and the additional set of probes.

In some cases, the additional set of probes recognizes one but no morethan ten genes. In some cases, the additional set of probes recognizes2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 genes selected fromIFNB1, IFNE, IFNW1, ADAR, IFIT, IFI, IRF, OAS1, IRAK1, TNFAIP3, ATG5,TYK2, STAT4, OPN, and KRT.

In some cases, the expression level of one or more genes selected fromIFNB1, IFNE, IFNW1, ADAR, IFIT, IFI, IRF, OAS1, IRAK1, TNFAIP3, ATG5,TYK2, STAT4, OPN, and KRT is an elevated gene expression level. In suchcases, the gene expression level is elevated by at least 1-fold, 2-fold,3-fold, 4-fold, 5-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold,60-fold, 70-fold, 80-fold, 90-fold, 100-fold, 110-fold, 120-fold,130-fold, 150-fold, 200-fold, 300-fold, 500-fold, or more. In someinstances, the gene expression level is elevated by at least 10%, 20%,30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, 300%, 400%, 500%, ormore. In some instances, the expression level is compared to a geneexpression level of an equivalent gene from a control sample. In someinstances, the control sample is a normal skin sample.

In some embodiments, a method described herein further comprisesdetecting a skin region affected with lupus erythematosus. In somecases, also described herein include a method monitoring the skin regionaffected with lupus erythematosus, for about 1 week, 2 weeks, 3 weeks, 1month, 2 months, 6 months, or more.

In some instances, the method has an improved specificity, of at leastor about 70%, 75%, 80%, 85%, 90%, or more than 95% when detecting thegene expression level of IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, CCL5, or acombination thereof. In some embodiments, the specificity is at least orabout 70%, 75%, 80%, 85%, 90%, or more than 95% when detecting the geneexpression level of IFNB1, IFNE, IFNW1, ADAR, IFIT, IFI, IRF, OAS1,IRAK1, TNFAIP3, ATG5, TYK2, STAT4, OPN, KRT, or a combination thereof.

In some cases, the method also has an improved sensitivity. In someembodiments, the sensitivity is at least or about 70%, 75%, 80%, 85%,90%, or more than 95% when detecting the gene expression levels ofIFNA1, IFNA2, IFNA4, IFNR1, IFNR2, CCL5, or a combination thereof. Insome cases, the sensitivity is at least or about 70%, 75%, 80%, 85%,90%, or more than 95% when detecting the gene expression levels ofIFNB1, IFNE, IFNW1, ADAR, IFIT, IFI, IRF, OAS1, IRAK1, TNFAIP3, ATG5,TYK2, STAT4, OPN, KRT, or a combination thereof.

In some embodiments, a method described herein comprises detecting geneexpression levels from a first gene classifier and a second geneclassifier in a subject in need thereof, comprising: (a) isolatingnucleic acids from a skin sample obtained from the subject, wherein theskin sample (e.g., comprising cells from the stratum corneum); (b)detecting the expression levels of one or more genes from the first geneclassifier: IFNA1, IFNA2, IFNA4, IFNR1, IFNR2, and CCL5, by contactingthe isolated nucleic acids with a set of probes that recognizes one ormore genes from the first gene classifier, and detects binding betweenone or more genes from the first gene classifier and the set of probes;and (c) detecting the expression levels of one or more genes from thesecond gene classifier: IFNB1, IFNE, IFNW1, ADAR, IFIT, IFI, IRF, OAS1,IRAK1, TNFAIP3, ATG5, TYK2, STAT4, OPN, and KRT, by contacting theisolated nucleic acids with an additional set of probes that recognizesone or more genes from the second gene classifier, and detects bindingbetween one or more genes from the second gene classifier and theadditional set of probes.

Current treatment forms of lupus are focused on induction therapy toachieve remission and long-term maintenance therapy to prevent relapse.Management of cutaneous forms of the disease can include treatment withantimalarials, dapsone, retinoids, corticosteroids, immunosuppressivedrugs, or thalidomide. In some instances, the animalarialhydroxychloroquine has been shown to decrease flares and assist inlong-term management of disease. A recent study has shown that Januskinase inhibitors (Jakinibs) are efficacious in improving diseasesymptoms in mice (Mok, C C, Expert Opin Investig Drugs. 2019 January;28(1):85-92).

Provided herein are methods for treatment of a subject determined tohave lupus by a non-inventive method described herein. Compositions forinclusion in methods for treatment of lupus described herein include,but are not limited to, antimalarials, dapsone, retinoids,corticosteroids, immunosuppressive drugs, thalidomide, Janus kinaseinhibitors, baricitinib or any combination thereof. In some cases, thecomposition for inclusion in the methods described herein for treatmentof lupus comprises an antimalarial. In some cases, the composition forinclusion in the methods described herein for treatment of lupuscomprises dapsone. In some cases, the composition for inclusion in themethods described herein for treatment of lupus comprises a retinoid. Insome cases, the composition for inclusion in the methods describedherein for treatment of lupus comprises a corticosteroid. In some cases,the composition for inclusion in the methods described herein fortreatment of lupus comprises an immunosuppressive drug. In some cases,the composition for inclusion in the methods described herein fortreatment of lupus comprises thalidomide. In some cases, the compositionfor inclusion in the methods described herein for treatment of lupuscomprises a Janus kinase inhibitor. In some cases, the composition forinclusion in the methods described herein for treatment of lupuscomprises baricitinib.

In some cases, the antimalarial is hydroxychloroquine. In some cases,the antimalarial is quinacrine. In some cases, the antimalarial ischloroquine.

In some cases, the immunosuppressive drug is methotrexate. In somecases, the immunosuppressive drug is azathioprine.

Diagnostic Tools and Methods

In some embodiments, one or more genes are detected with a set ofprobes. In some embodiments, the set of probes comprises at least orabout 1, 2, 3, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19,20, 25, 30, or more than 30 probes. In some embodiments, the set ofprobes comprises about 6 probes. In some embodiments, the set of probescomprises about 7 probes. In some embodiments, the set of probescomprises about 8 probes. In some embodiments, the set of probescomprises about 9 probes. In some embodiments, the set of probescomprises about 10 probes. In some embodiments, the set of probescomprises about 13 probes. In some embodiments, the set of probescomprises about 15 probes. In some embodiments, the set of probescomprises about 20 probes.

In some embodiments, the set of probes comprises one or more primerpairs. In some embodiments, a number of primer pairs is at least orabout 1, 2, 3, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19,20, 25, 30, or more than 30 primer pairs. In some embodiments, thenumber of primer pairs is about 8 primer pairs. In some embodiments, thenumber of primer pairs is about 9 primer pairs. In some embodiments, thenumber of primer pairs is about 10 primer pairs.

In some embodiments, one or more probes in the set of probes is labeled.In some embodiments, the one or more probe is labeled with a radioactivelabel, a fluorescent label, an enzyme, a chemiluminescent tag, acolorimetric tag, an affinity tag or other labels or tags that are knownin the art.

Exemplary affinity tags include, but are not limited to, biotin,desthiobiotin, histidine, polyhistidine, myc, hemagglutinin (HA), FLAG,glutathione S transferase (GST), or derivatives thereof. In someembodiments, the affinity tag is recognized by avidin, streptavidin,nickel, or glutathione.

In some embodiments, the fluorescent label is a fluorophore, afluorescent protein, a fluorescent peptide, quantum dots, a fluorescentdye, a fluorescent material, or variations or combinations thereof.

Exemplary fluorophores include, but are not limited to, Alexa-Fluor dyes(e.g., Alexa Fluor® 350, Alexa Fluor® 405, Alexa Fluor® 430, AlexaFluor® 488, Alexa Fluor® 500, Alexa Fluor® 514, Alexa Fluor® 532, AlexaFluor® 546, Alexa Fluor® 555, Alexa Fluor® 568, Alexa Fluor® 594, AlexaFluor® 610, Alexa Fluor® 633, Alexa Fluor® 647, Alexa Fluor® 660, AlexaFluor® 680, Alexa Fluor® 700, and Alexa Fluor® 750), APC, Cascade Blue,Cascade Yellow and R-phycoerythrin (PE), DyLight 405, DyLight 488,DyLight 550, DyLight 650, DyLight 680, DyLight 755, DyLight 800, FITC,Pacific Blue, PerCP, Rhodamine, and Texas Red, Cy5, Cy5.5, Cy7.

Examples of fluorescent peptides include GFP (Green Fluorescent Protein)or derivatives of GFP (e.g., EBFP, EBFP2, Azurite, mKalamal, ECFP,Cerulean, CyPet, YFP, Citrine, Venus, and YPet.

Examples of fluorescent dyes include, but are not limited to, xanthenes(e.g., rhodamines, rhodols and fluoresceins, and their derivatives);bimanes; coumarins and their derivatives (e.g., umbelliferone andaminomethyl coumarins); aromatic amines (e.g., dansyl; squarate dyes);benzofurans; fluorescent cyanines; indocarbocyanines; carbazoles;dicyanomethylene pyranes; polymethine; oxabenzanthrane; xanthene;pyrylium; carbostyl; perylene; acridone; quinacridone; rubrene;anthracene; coronene; phenanthrecene; pyrene; butadiene; stilbene;porphyrin; pthalocyanine; lanthanide metal chelate complexes; rare-earthmetal chelate complexes; and derivatives of such dyes. In someembodiments, the fluorescein dye is, but not limited to,5-carboxyfluorescein, fluorescein-5-isothiocyanate,fluorescein-6-isothiocyanate and 6-carboxyfluorescein. In someembodiments, the rhodamine dye is, but not limited to,tetramethylrhodamine-6-isothiocyanate, 5-carboxytetramethylrhodamine,5-carboxy rhodol derivatives, tetramethyl and tetraethyl rhodamine,diphenyldimethyl and diphenyldiethyl rhodamine, dinaphthyl rhodamine,and rhodamine 101 sulfonyl chloride (sold under the tradename of TEXASRED®). In some embodiments, the cyanine dye is Cy3, Cy3B, Cy3.5, Cy5,Cy5.5, Cy7, IRDYE680, Alexa Fluor 750, IRDye800CW, or ICG.

In some embodiments, the gene expression levels of IL-13, TSLP, IL-31,or a combination thereof is measured using PCR. Examples of PCRtechniques include, but are not limited to quantitative PCR (qPCR),single cell PCR, PCR-RFLP, digital PCR (dPCR), droplet digital PCR(ddPCR), single marker qPCR, hot start PCR, and Nested PCR.

In some embodiments, the gene expression levels of IL-13R, IL-4R, IL-17,IL-22, CXCL9, CXCL10, CXCL11, S100A7, S100A8, S100A9, CCL17, CCL18,CCL19, CCL26, CCL27, NOS2, or a combination thereof is measured usingPCR. Examples of PCR techniques include, but are not limited toquantitative PCR (qPCR), single cell PCR, PCR-RFLP, digital PCR (dPCR),droplet digital PCR (ddPCR), single marker qPCR, hot start PCR, andNested PCR.

In some embodiments, the gene expression levels of IL-17A, IL-17F, IL-8,CXCL5, S100A9, DEFB4A, or a combination thereof is measured using PCR.Examples of PCR techniques include, but are not limited to quantitativePCR (qPCR), single cell PCR, PCR-RFLP, digital PCR (dPCR), dropletdigital PCR (ddPCR), single marker qPCR, hot start PCR, and Nested PCR.

In some embodiments, the gene expression levels of IL-17C, S100A7,IL-17RA, IL-17RC, IL-23A, IL-22, IL-6, IL-24, IL-6, CXCL1, IFN-gamma,IL-31, IL-33, TNFα, LCN2, CCL20, and TNFRSF1A, or a combination thereofis measured using PCR. Examples of PCR techniques include, but are notlimited to quantitative PCR (qPCR), single cell PCR, PCR-RFLP, digitalPCR (dPCR), droplet digital PCR (ddPCR), single marker qPCR, hot startPCR, and Nested PCR.

In some embodiments, the gene expression levels of IFNA1, IFNA2, IFNA4,IFNR1, IFNR2, CCL5, or a combination thereof is measured using PCR.Examples of PCR techniques include, but are not limited to quantitativePCR (qPCR), single cell PCR, PCR-RFLP, digital PCR (dPCR), dropletdigital PCR (ddPCR), single marker qPCR, hot start PCR, and Nested PCR.

In some embodiments, the gene expression levels of IFNB1, IFNE, IFNW1,ADAR, IFIT, IFI, IRF, OAS1, IRAK1, TNFAIP3, ATG5, TYK2, STAT4, OPN, KRT,or a combination thereof is measured using PCR. Examples of PCRtechniques include, but are not limited to quantitative PCR (qPCR),single cell PCR, PCR-RFLP, digital PCR (dPCR), droplet digital PCR(ddPCR), single marker qPCR, hot start PCR, and Nested PCR.

In some embodiments, the expression levels are measured using qPCR. Insome embodiments, the qPCR comprises use of fluorescent dyes orfluorescent probes. In some embodiments, the fluorescent dye is anintercalating dye. Examples of intercalating dyes include, but are notlimited to, intercalating dyes include SYBR green I, SYBR green II, SYBRgold, ethidium bromide, methylene blue, Pyronin Y, DAPI, acridineorange, Blue View, or phycoerythrin. In some embodiments, the qPCRcomprises use of more than one fluorescent probe. In some embodiments,the use of more than one fluorescent probes allows for multiplexing. Forexample, different non-classical variants are hybridized to differentfluorescent probes and can be detected in a single qPCR reaction.

Components of the Skin Collection Kit

In some embodiments, the adhesive patch from the sample collection kitdescribed herein comprises a first collection area comprising anadhesive matrix and a second area extending from the periphery of thefirst collection area. The adhesive matrix is located on a skin facingsurface of the first collection area. The second area functions as atab, suitable for applying and removing the adhesive patch. The tab issufficient in size so that while applying the adhesive patch to a skinsurface, the applicant does not come in contact with the matrix materialof the first collection area. In some embodiments, the adhesive patchdoes not contain a second area tab. In some instances, the adhesivepatch is handled with gloves to reduce contamination of the adhesivematrix prior to use.

In some embodiments, the first collection area is a polyurethane carrierfilm. In some embodiments, the adhesive matrix is comprised of asynthetic rubber compound. In some embodiments, the adhesive matrix is astyrene-isoprene-styrene (SIS) linear block copolymer compound. In someinstances, the adhesive patch does not comprise latex, silicone, orboth. In some instances, the adhesive patch is manufactured by applyingan adhesive material as a liquid-solvent mixture to the first collectionarea and subsequently removing the solvent.

The matrix material is sufficiently sticky to adhere to a skin sample.The matrix material is not so sticky that is causes scarring or bleedingor is difficult to remove. In some embodiments, the matrix material iscomprised of a transparent material. In some instances, the matrixmaterial is biocompatible. In some instances, the matrix material doesnot leave residue on the surface of the skin after removal. In certaininstances, the matrix material is not a skin irritant.

In some embodiments, the adhesive patch comprises a flexible material,enabling the patch to conform to the shape of the skin surface uponapplication. In some instances, at least the first collection area isflexible. In some instances, the tab is plastic. In an illustrativeexample, the adhesive patch does not contain latex, silicone, or both.In some embodiments, the adhesive patch is made of a transparentmaterial, so that the skin sampling area of the subject is visible afterapplication of the adhesive patch to the skin surface. The transparencyensures that the adhesive patch is applied on the desired area of skincomprising the skin area to be sampled. In some embodiments, theadhesive patch is between about 5 and about 100 mm in length. In someembodiments, the first collection area is between about 5 and about 40mm in length. In some embodiments, the first collection area is betweenabout 10 and about 20 mm in length. In some embodiments the length ofthe first collection area is configured to accommodate the area of theskin surface to be sampled, including, but not limited to, about 19 mm,about 20 mm, about 21 mm, about 22 mm, about 23 mm, about 24 mm, about25 mm, about 30 mm, about 35 mm, about 40 mm, about 45 mm, about 50 mm,about 55 mm, about 60 mm, about 65 mm, about 70 mm, about 75 mm, about80 mm, about 85 mm, about 90 mm, and about 100 mm. In some embodiments,the first collection area is elliptical.

In further embodiments, the adhesive patch of this invention is providedon a peelable release sheet in the adhesive skin sample collection kit.In some embodiments, the adhesive patch provided on the peelable releasesheet is configured to be stable at temperatures between −80° C. and 30°C. for at least 6 months, at least 1 year, at least 2 years, at least 3years, and at least 4 years. In some instances, the peelable releasesheet is a panel of a tri-fold skin sample collector.

In some instances, nucleic acids are stable on adhesive patch or patcheswhen stored for a period of time or at a particular temperature. In someinstances, the period of time is at least or about 1 day, 2 days, 3days, 4 days, 5 days, 6 days, 7 days, 2 weeks, 3 weeks, 4 weeks, or morethan 4 weeks. In some instances, the period of time is about 7 days. Insome instances, the period of time is about 10 days. In some instances,the temperature is at least or about −80° C., −70° C., −60° C., −50° C.,−40° C., −20° C., −10° C., −4° C., 0° C., 5° C., 15° C., 18° C., 20° C.,25° C., 30° C., 35° C., 40° C., 45° C., 50° C., or more than 50° C. Thenucleic acids on the adhesive patch or patches, in some embodiments, arestored for any period of time described herein and any particulartemperature described herein. For example, the nucleic acids on theadhesive patch or patches are stored for at least or about 7 days atabout 25° C., 7 days at about 30° C., 7 days at about 40° C., 7 days atabout 50° C., 7 days at about 60° C., or 7 days at about 70° C. In someinstances, the nucleic acids on the adhesive patch or patches are storedfor at least or about 10 days at about −80° C.

The peelable release sheet, in certain embodiments, is configured tohold a plurality of adhesive patches, including, but not limited to, 12,11, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, from about 2 to about 8, from about 2to about 7, from about 2 to about 6, from about 2 to about 4, from about3 to about 6, from about 3 to about 8, from about 4 to about 10, fromabout 4 to about 8, from about 4 to about 6, from about 4 to about 5,from about 6 to about 10, from about 6 to about 8, or from about 4 toabout 8. In some instances, the peelable release sheet is configured tohold about 12 adhesive patches. In some instances, the peelable releasesheet is configured to hold about 11 adhesive patches. In someinstances, the peelable release sheet is configured to hold about 10adhesive patches. In some instances, the peelable release sheet isconfigured to hold about 9 adhesive patches. In some instances, thepeelable release sheet is configured to hold about 8 adhesive patches.In some instances, the peelable release sheet is configured to holdabout 7 adhesive patches. In some instances, the peelable release sheetis configured to hold about 6 adhesive patches. In some instances, thepeelable release sheet is configured to hold about 5 adhesive patches.In some instances, the peelable release sheet is configured to holdabout 4 adhesive patches. In some instances, the peelable release sheetis configured to hold about 3 adhesive patches. In some instances, thepeelable release sheet is configured to hold about 2 adhesive patches.In some instances, the peelable release sheet is configured to holdabout 1 adhesive patch.

Provided herein, in certain embodiments, are methods and compositionsfor obtaining a sample using an adhesive patch, wherein the adhesivepatch is applied to the skin and removed from the skin. After removingthe used adhesive patch from the skin surface, the patch strippingmethod, in some instances, further comprise storing the used patch on aplacement area sheet, where the patch remains until the skin sample isisolated or otherwise utilized. In some instances, the used patch isconfigured to be stored on the placement area sheet for at least 1 weekat temperatures between −80° C. and 30° C. In some embodiments, the usedpatch is configured to be stored on the placement area sheet for atleast 2 weeks, at least 3 weeks, at least 1 month, at least 2 months, atleast 3 months, at least 4 months, at least 5 months, and at least 6months at temperatures between −80° C. to 30° C.

In some instances, the placement area sheet comprises a removable liner,provided that prior to storing the used patch on the placement areasheet, the removable liner is removed. In some instances, the placementarea sheet is configured to hold a plurality of adhesive patches,including, but not limited to, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1,from about 2 to about 8, from about 2 to about 7, from about 2 to about6, from about 2 to about 4, from about 3 to about 6, from about 3 toabout 8, from about 4 to about 10, from about 4 to about 8, from about 4to about 6, from about 4 to about 5, from about 6 to about 10, fromabout 6 to about 8, or from about 4 to about 8. In some instances, theplacement area sheet is configured to hold about 12 adhesive patches. Insome instances, the placement area sheet is configured to hold about 11adhesive patches. In some instances, the placement area sheet isconfigured to hold about 10 adhesive patches. In some instances, theplacement area sheet is configured to hold about 9 adhesive patches. Insome instances, the placement area sheet is configured to hold about 8adhesive patches. In some instances, the placement area sheet isconfigured to hold about 7 adhesive patches. In some instances, theplacement area sheet is configured to hold about 6 adhesive patches. Insome instances, the placement area sheet is configured to hold about 5adhesive patches. In some instances, the placement area sheet isconfigured to hold about 4 adhesive patches. In some instances, theplacement area sheet is configured to hold about 3 adhesive patches. Insome instances, the placement area sheet is configured to hold about 2adhesive patches. In some instances, the placement area sheet isconfigured to hold about 1 adhesive patch.

The used patch, in some instances, is stored so that the matrixcontaining, skin facing surface of the used patch is in contact with theplacement area sheet. In some instances, the placement area sheet is apanel of the tri-fold skin sample collector. In some instances, thetri-fold skin sample collector further comprises a clear panel. In someinstances, the tri-fold skin sample collector is labeled with a uniquebarcode that is assigned to a subject. In some instances, the tri-foldskin sample collector comprises an area for labeling subjectinformation.

In an illustrative embodiment, the adhesive skin sample collection kitcomprises the tri-fold skin sample collector comprising adhesive patchesstored on a peelable release panel. In some instances, the tri-fold skinsample collector further comprises a placement area panel with aremovable liner. In some instances, the patch stripping method involvesremoving an adhesive patch from the tri-fold skin sample collectorpeelable release panel, applying the adhesive patch to a skin sample,removing the used adhesive patch containing a skin sample and placingthe used patch on the placement area sheet. In some instances, theplacement area panel is a single placement area panel sheet. In someinstances, the identity of the skin sample collected is indexed to thetri-fold skin sample collector or placement area panel sheet by using abarcode or printing patient information on the collector or panel sheet.In some instances, the indexed tri-fold skin sample collector orplacement sheet is sent to a diagnostic lab for processing. In someinstances, the used patch is configured to be stored on the placementpanel for at least 1 week at temperatures between −80° C. and 25° C. Insome embodiments, the used patch is configured to be stored on theplacement area panel for at least 2 weeks, at least 3 weeks, at least 1month, at least 2 months, at least 3 months, at least 4 months, at least5 months, and at least 6 months at temperatures between −80° C. and 25°C. In some embodiments, the indexed tri-fold skin sample collector orplacement sheet is sent to a diagnostic lab using UPS or FedEx.

In an exemplary embodiment, the patch stripping method further comprisespreparing the skin sample prior to application of the adhesive patch.Preparation of the skin sample includes, but is not limited to, removinghairs on the skin surface, cleansing the skin surface and/or drying theskin surface. In some instances, the skin surface is cleansed with anantiseptic including, but not limited to, alcohols, quaternary ammoniumcompounds, peroxides, chlorhexidine, halogenated phenol derivatives andquinolone derivatives. In some instances, the alcohol is about 0 toabout 20%, about 20 to about 40%, about 40 to about 60%, about 60 toabout 80%, or about 80 to about 100% isopropyl alcohol. In someinstances, the antiseptic is 70% isopropyl alcohol.

In some embodiments, the patch stripping method is used to collect askin sample from the surfaces including, but not limited to, the face,head, neck, arm, chest, abdomen, back, leg, hand or foot. In someinstances, the skin surface is not located on a mucous membrane. In someinstances, the skin surface is not ulcerated or bleeding. In certaininstances, the skin surface has not been previously biopsied. In certaininstances, the skin surface is not located on the soles of the feet orpalms.

The patch stripping method, devices, and systems described herein areuseful for the collection of a skin sample from a skin lesion. A skinlesion is a part of the skin that has an appearance or growth differentfrom the surrounding skin. In some instances, the skin lesion ispigmented. A pigmented lesion includes, but is not limited to, a mole,dark colored skin spot and a melanin containing skin area. In someembodiments, the skin lesion is from about 5 mm to about 16 mm indiameter. In some instances, the skin lesion is from about 5 mm to about15 mm, from about 5 mm to about 14 mm, from about 5 mm to about 13 mm,from about 5 mm to about 12 mm, from about 5 mm to about 11 mm, fromabout 5 mm to about 10 mm, from about 5 mm to about 9 mm, from about 5mm to about 8 mm, from about 5 mm to about 7 mm, from about 5 mm toabout 6 mm, from about 6 mm to about 15 mm, from about 7 mm to about 15mm, from about 8 mm to about 15 mm, from about 9 mm to about 15 mm, fromabout 10 mm to about 15 mm, from about 11 mm to about 15 mm, from about12 mm to about 15 mm, from about 13 mm to about 15 mm, from about 14 mmto about 15 mm, from about 6 to about 14 mm, from about 7 to about 13mm, from about 8 to about 12 mm and from about 9 to about 11 mm indiameter. In some embodiments, the skin lesion is from about 10 mm toabout 20 mm, from about 20 mm to about 30 mm, from about 30 mm to about40 mm, from about 40 mm to about 50 mm, from about 50 mm to about 60 mm,from about 60 mm to about 70 mm, from about 70 mm to about 80 mm, fromabout 80 mm to about 90 mm, and from about 90 mm to about 100 mm indiameter. In some instances, the diameter is the longest diameter of theskin lesion. In some instances, the diameter is the smallest diameter ofthe skin lesion.

The adhesive skin sample collection kit, in some embodiments, comprisesat least one adhesive patch, a sample collector, and an instruction foruse sheet. In an exemplary embodiment, the sample collector is atri-fold skin sample collector comprising a peelable release panelcomprising at least one adhesive patch, a placement area panelcomprising a removable liner, and a clear panel. The tri-fold skinsample collector, in some instances, further comprises a barcode and/oran area for transcribing patient information. In some instances, theadhesive skin sample collection kit is configured to include a pluralityof adhesive patches, including but not limited to 12, 11, 10, 9, 8, 7,6, 5, 4, 3, 2, 1, from about 2 to about 8, from about 2 to about 7, fromabout 2 to about 6, from about 2 to about 4, from about 3 to about 6,from about 3 to about 8, from about 4 to about 10, from about 4 to about8, from about 4 to about 6, from about 4 to about 5, from about 6 toabout 10, from about 6 to about 8, or from about 4 to about 8. Theinstructions for use sheet provide the kit operator all of the necessaryinformation for carrying out the patch stripping method. Theinstructions for use sheet preferably include diagrams to illustrate thepatch stripping method.

In some instances, the adhesive skin sample collection kit provides allthe necessary components for performing the patch stripping method. Insome embodiments, the adhesive skin sample collection kit includes a labrequisition form for providing patient information. In some instances,the kit further comprises accessory components. Accessory componentsinclude, but are not limited to, a marker, a resealable plastic bag,gloves and a cleansing reagent. The cleansing reagent includes, but isnot limited to, an antiseptic such as isopropyl alcohol. In someinstances, the components of the skin sample collection kit are providedin a cardboard box.

Cellular Material and Sample Process

The methods and devices provided herein, in certain embodiments, involveapplying an adhesive or other similar patch to the skin in a manner sothat an effective or sufficient amount of a tissue, such as a skinsample, adheres to the adhesive matrix of the adhesive patch. Forexample, the effective or sufficient amount of a skin sample is anamount that removably adheres to a material, such as the matrix oradhesive patch. The adhered skin sample, in certain embodiments,comprises cellular material including nucleic acids. In some instances,the nucleic acid is RNA or DNA. An effective amount of a skin samplecontains an amount of cellular material sufficient for performing adiagnostic assay. In some instances, the diagnostic assay is performedusing the cellular material isolated from the adhered skin sample on theused adhesive patch. In some instances, the diagnostic assay isperformed on the cellular material adhered to the used adhesive patch.In some embodiments, an effect amount of a skin sample comprises anamount of RNA sufficient to perform a gene expression analysis.Sufficient amounts of RNA includes, but not limited to, picogram,nanogram, and microgram quantities.

In some instances, the nucleic acid is a RNA molecule or a fragmentedRNA molecule (RNA fragments). In some instances, the RNA is a microRNA(miRNA), a pre-miRNA, a pri-miRNA, a mRNA, a pre-mRNA, a viral RNA, aviroid RNA, a virusoid RNA, circular RNA (circRNA), a ribosomal RNA(rRNA), a transfer RNA (tRNA), a pre-tRNA, a long non-coding RNA(lncRNA), a small nuclear RNA (snRNA), a circulating RNA, a cell-freeRNA, an exosomal RNA, a vector-expressed RNA, a RNA transcript, asynthetic RNA, or combinations thereof. In some instances, the RNA ismRNA. In some instances, the RNA is cell-free circulating RNA.

In some instances, the nucleic acid is DNA. DNA includes, but notlimited to, genomic DNA, viral DNA, mitochondrial DNA, plasmid DNA,amplified DNA, circular DNA, circulating DNA, cell-free DNA, or exosomalDNA. In some instances, the DNA is single-stranded DNA (ssDNA),double-stranded DNA, denaturing double-stranded DNA, synthetic DNA, andcombinations thereof. In some instances, the DNA is genomic DNA. In someinstances, the DNA is cell-free circulating DNA.

In additional embodiments, the adhered skin sample comprises cellularmaterial including nucleic acids such as RNA or DNA, in an amount thatis at least about 1 picogram. In some embodiments, the amount ofcellular material is no more than about 1 nanogram. In further oradditional embodiments, the amount of cellular material is no more thanabout 1 microgram. In still further or additional embodiments, theamount of cellular material is no more than about 1 gram.

In further or additional embodiments, the amount of cellular material isfrom about 1 picogram to about 1 gram. In further or additionalembodiments, the cellular material comprises an amount that is fromabout 50 microgram to about 1 gram, from about 100 picograms to about500 micrograms, from about 500 picograms to about 100 micrograms, fromabout 750 picograms to about 1 microgram, from about 1 nanogram to about750 nanograms, or from about 1 nanogram to about 500 nanograms. Inadditional embodiments, the cellular material comprises an amount thatis from about 50 picograms to about 1 micrograms, from about 100picograms to about 500 picograms, from about 200 picograms to about 500picograms, from about 500 picograms to about 1 nanograms, from about 500picograms to about 500 nanograms, or from about 1 nanograms to about 500nanograms.

In further or additional embodiments, the amount of cellular material,including nucleic acids such as RNA or DNA, comprises an amount that isfrom about 50 microgram to about 500 microgram, from about 100 microgramto about 450 microgram, from about 100 microgram to about 350 microgram,from about 100 microgram to about 300 microgram, from about 120microgram to about 250 microgram, from about 150 microgram to about 200microgram, from about 500 nanograms to about 5 nanograms, or from about400 nanograms to about 10 nanograms, or from about 200 nanograms toabout 15 nanograms, or from about 100 nanograms to about 20 nanograms,or from about 50 nanograms to about 10 nanograms, or from about 50nanograms to about 25 nanograms. In some embodiments, the amount ofcellular material, including nucleic acids such as RNA or DNA, comprisesan amount that is from about picograms to about 1 micrograms, from about100 picograms to about 500 picograms, from about 200 picograms to about500 picograms, from about 500 picograms to about 1 nanograms, from about500 picograms to about 500 nanograms, or from about 1 nanograms to about500 nanograms.

In further or additional embodiments, the amount of cellular material,including nucleic acids such as RNA or DNA, is less than about 1 gram,is less than about 500 micrograms, is less than about 490 micrograms, isless than about 480 micrograms, is less than about 470 micrograms, isless than about 460 micrograms, is less than about 450 micrograms, isless than about 440 micrograms, is less than about 430 micrograms, isless than about 420 micrograms, is less than about 410 micrograms, isless than about 400 micrograms, is less than about 390 micrograms, isless than about 380 micrograms, is less than about 370 micrograms, isless than about 360 micrograms, is less than about 350 micrograms, isless than about 340 micrograms, is less than about 330 micrograms, isless than about 320 micrograms, is less than about 310 micrograms, isless than about 300 micrograms, is less than about 290 micrograms, isless than about 280 micrograms, is less than about 270 micrograms, isless than about 260 micrograms, is less than about 250 micrograms, isless than about 240 micrograms, is less than about 230 micrograms, isless than about 220 micrograms, is less than about 210 micrograms, isless than about 200 micrograms, is less than about 190 micrograms, isless than about 180 micrograms, is less than about 170 micrograms, isless than about 160 micrograms, is less than about 150 micrograms, isless than about 140 micrograms, is less than about 130 micrograms, isless than about 120 micrograms, is less than about 110 micrograms, isless than about 100 micrograms, is less than about 90 micrograms, isless than about 80 micrograms, is less than about 70 micrograms, is lessthan about 60 micrograms, is less than about 50 micrograms, is less thanabout 20 micrograms, is less than about 10 micrograms, is less thanabout 5 micrograms, is less than about 1 microgram, is less than about750 nanograms, is less than about 500 nanograms, is less than about 250nanograms, is less than about 150 nanograms, is less than about 100nanograms, is less than about 50 nanograms, is less than about 25nanograms, is less than about 15 nanograms, is less than about 1nanogram, is less than about 750 picograms, is less than about 500picograms, is less than about 250 picograms, is less than about 100picograms, is less than about 50 picograms, is less than about 25picograms, is less than about 15 picograms, or is less than about 1picogram.

In some embodiments, isolated RNA from a collected skin sample isreverse transcribed into cDNA, for example for amplification by PCR toenrich for target genes. The expression levels of these target genes arequantified by quantitative PCR in a gene expression test. In someinstances, in combination with quantitative PCR, a software programperformed on a computer is utilized to quantify RNA isolated from thecollected skin sample. In some instances, a software program or moduleis utilized to relate a quantity of RNA from a skin sample to a geneexpression signature, wherein the gene expression signature isassociated with a disease such as skin cancer. In some embodiments, asoftware program or module scores a sample based on gene expressionlevels. In some embodiments, the sample score is compared with areference sample score to determine if there is a statisticalsignificance between the gene expression signature and a disease.

In some instances, the layers of skin include epidermis, dermis, orhypodermis. The outer layer of epidermis is the stratum corneum layer,followed by stratum lucidum, stratum granulosum, stratum spinosum, andstratum basale. In some instances, the skin sample is obtained from theepidermis layer. In some cases, the skin sample is obtained from thestratum corneum layer. In some instances, the skin sample is obtainedfrom the dermis.

In some instances, cells from the stratum corneum layer are obtained,which comprises keratinocytes. In some cases, melanocytes are notobtained from the skin sample.

Following extraction of nucleic acids from a biological sample, thenucleic acids, in some instances, are further purified. In someinstances, the nucleic acids are RNA. In some instances, the nucleicacids are DNA. In some instances, the RNA is human RNA. In someinstances, the DNA is human DNA. In some instances, the RNA is microbialRNA. In some instances, the DNA is microbial DNA. In some instances,human nucleic acids and microbial nucleic acids are purified from thesame biological sample. In some instances, nucleic acids are purifiedusing a column or resin based nucleic acid purification scheme. In someinstances, this technique utilizes a support comprising a surface areafor binding the nucleic acids. In some instances, the support is made ofglass, silica, latex or a polymeric material. In some instances, thesupport comprises spherical beads.

Methods for isolating nucleic acids, in certain embodiments, compriseusing spherical beads. In some instances, the beads comprise materialfor isolation of nucleic acids. Exemplary material for isolation ofnucleic acids using beads include, but not limited to, glass, silica,latex, and a polymeric material. In some instances, the beads aremagnetic. In some instances, the beads are silica coated. In someinstances, the beads are silica-coated magnetic beads. In someinstances, a diameter of the spherical bead is at least or about 0.5 um,1 um, 1.5 um, 2 um, 2.5 um, 3 um, 3.5 um, 4 um, 4.5 um, 5 um, 5.5 um, 6um, 6.5 um, 7 um, 7.5 um, 8 um, 8.5 um, 9 um, 9.5 um, 10 um, or morethan 10 um.

In some cases, a yield of the nucleic acids products obtained usingmethods described herein is about 500 picograms or higher, about 600picograms or higher, about 1000 picograms or higher, about 2000picograms or higher, about 3000 picograms or higher, about 4000picograms or higher, about 5000 picograms or higher, about 6000picograms or higher, about 7000 picograms or higher, about 8000picograms or higher, about 9000 picograms or higher, about 10000picograms or higher, about 20000 picograms or higher, about 30000picograms or higher, about 40000 picograms or higher, about 50000picograms or higher, about 60000 picograms or higher, about 70000picograms or higher, about 80000 picograms or higher, about 90000picograms or higher, or about 100000 picograms or higher.

In some cases, a yield of the nucleic acids products obtained usingmethods described herein is about 100 picograms, 500 picograms, 600picograms, 700 picograms, 800 picograms, 900 picograms, 1 nanogram, 5nanograms, 10 nanograms, 15 nanograms, 20 nanograms, 21 nanograms, 22nanograms, 23 nanograms, 24 nanograms, 25 nanograms, 26 nanograms, 27nanograms, 28 nanograms, 29 nanograms, 30 nanograms, 35 nanograms, 40nanograms, 50 nanograms, 60 nanograms, 70 nanograms, 80 nanograms, 90nanograms, 100 nanograms, 500 nanograms, or higher.

In some cases, methods described herein provide less than less than 10%,less than 8%, less than 5%, less than 2%, less than 1%, or less than0.5% product yield variations between samples.

In some cases, methods described herein provide a substantiallyhomogenous population of a nucleic acid product.

In some cases, methods described herein provide less than 30%, less than25%, less than 20%, less than 15%, less than 10%, less than 8%, lessthan 5%, less than 2%, less than 1%, or less than 0.5% contaminants.

In some instances, following extraction, nucleic acids are stored. Insome instances, the nucleic acids are stored in water, Tris buffer, orTris-EDTA buffer before subsequent analysis. In some instances, thisstorage is less than 8° C. In some instances, this storage is less than4° C. In certain embodiments, this storage is less than 0° C. In someinstances, this storage is less than −20° C. In certain embodiments,this storage is less than −70° C. In some instances, the nucleic acidsare stored for about 1, 2, 3, 4, 5, 6, or 7 days. In some instances, thenucleic acids are stored for about 1, 2, 3, or 4 weeks. In someinstances, the nucleic acids are stored for about 1, 2, 3, 4, 5, 6, 7,8, 9, 10, 11, or 12 months.

In some instances, nucleic acids isolated using methods described hereinare subjected to an amplification reaction following isolation andpurification. In some instances, the nucleic acids to be amplified areRNA including, but not limited to, human RNA and human microbial RNA. Insome instances, the nucleic acids to be amplified are DNA including, butnot limited to, human DNA and human microbial DNA. Non-limitingamplification reactions include, but are not limited to, quantitativePCR (qPCR), self-sustained sequence replication, transcriptionalamplification system, Q-Beta Replicase, rolling circle replication, orany other nucleic acid amplification known in the art. In someinstances, the amplification reaction is PCR. In some instances, theamplification reaction is quantitative such as qPCR.

Provided herein are methods for detecting an expression level of one ormore genes of interest from nucleic acids isolated from a biologicalsample. In some instances, the expression level is detected following anamplification reaction. In some instances, the nucleic acids are RNA. Insome instances, the RNA is human RNA. In some instances, the RNA ismicrobial RNA. In some instances, the nucleic acids are DNA. In someinstances, the DNA is human DNA. In some instances, the DNA is microbialDNA. In some instances, the expression level is determined using PCR. Insome instances, the expression level is determined using qPCR. In someinstances, the expression level is determined using a microarray. Insome instances, the expression level is determined by sequencing.

Certain Terminologies

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of skill in theart to which the claimed subject matter belongs. It is to be understoodthat the detailed description are exemplary and explanatory only and arenot restrictive of any subject matter claimed. In this application, theuse of the singular includes the plural unless specifically statedotherwise. It must be noted that, as used in the specification, thesingular forms “a,” “an” and “the” include plural referents unless thecontext clearly dictates otherwise. In this application, the use of “or”means “and/or” unless stated otherwise. Furthermore, use of the term“including” as well as other forms, such as “include”, “includes,” and“included,” is not limiting.

Although various features of the invention may be described in thecontext of a single embodiment, the features may also be providedseparately or in any suitable combination. Conversely, although theinvention may be described herein in the context of separate embodimentsfor clarity, the invention may also be implemented in a singleembodiment.

Reference in the specification to “some embodiments”, “an embodiment”,“one embodiment” or “other embodiments” means that a particular feature,structure, or characteristic described in connection with theembodiments is included in at least some embodiments, but notnecessarily all embodiments, of the inventions.

As used herein, ranges and amounts can be expressed as “about” aparticular value or range. About also includes the exact amount. Hence“about 5 μL” means “about 5 μL” and also “5 μL” Generally, the term“about” includes an amount that would be expected to be withinexperimental error.

The section headings used herein are for organizational purposes onlyand are not to be construed as limiting the subject matter described.

As used herein, the terms “individual(s)”, “subject(s)” and “patient(s)”mean any mammal. In some embodiments, the mammal is a human. In someembodiments, the mammal is a non-human. None of the terms require or arelimited to situations characterized by the supervision (e.g. constant orintermittent) of a health care worker (e.g. a doctor, a registerednurse, a nurse practitioner, a physician's assistant, an orderly or ahospice worker).

Markers

C-C Motif Chemokine Ligand 20 (CCL20), also known as small InducibleCytokine Subfamily A (Cys-Cys), Member 20, MIP3A, or ST38, is a memberof a family of small cytokine CC genes, the products of which arecharacterized by two adjacent cysteines. CCL20 encodes a protein withchemotactic activity for lymphocytes which can repress proliferation ofmyeloid progenitors. In some instances, CCL20 has Gene ID: 6364.

C-X-C Motif Chemokine Ligand 1 (CXCL1), also known as Chemokine (C-X-CMotif) Ligand 1, Fibroblast Secretory Protein, or GRO1, encodes aprotein in the CXC subfamily of chemokines. CXCL1 encodes a secretedgrowth factor that signals through the G-protein coupled receptor, CXCreceptor 2. In some instances, CXCL1 has Gene ID: 2919.

C-X-C Motif Chemokine Ligand 5 (CXCL5), also known as Small InducibleCytokine Subfamily B (Cys-X-Cys), Member 5, ENA-78, or SCYB5, encodes aprotein in the CXC subfamily of chemokines. The encoded protein isproposed to bind the G-protein coupled receptor chemokine (C-X-C motif)receptor 2 to recruit neutrophils, to promote angiogenesis, and toremodel connective tissues. In some instances, CXCL5 has Gene ID: 6374.

Defensin Beta 4A (DEFB4A), also known as Defensin Beta 4, HBD-2, orSAP1, is part of a family of microbicidal and cytotoxic peptides made byneutrophils. The encoded protein, defensin, beta 4, is an antibioticpeptide which is locally regulated by inflammation. In some instances,DEFB4A has Gene ID: 1673.

Interfereon Gamma (IFN-gamma), also known as IFN-gamma, or ImmuneInterferon, is part of the type II interferon class. The encoded proteinis a homodimer that binds to the interferon gamma receptor to trigger acellular response to viral and microbial infections. In some instances,IFNG has Gene ID: 3458.

Interleukin 17A (IL-17A), also known as IL-17, CTLA-8 or CytotoxicT-Lymphocyte-Associated Protein 8, is a proinflammatory cytokineproduced by activated T cells. This cytokine regulates the activities ofNF-kappaB and mitogen-activated protein kinases and can stimulate theexpression of IL6 and cyclooxygenase-2 (PTGS2/COX-2), as well as enhancethe production of nitric oxide (NO). In some instances, IL-17A has GeneID: 3605.

Interleukin 17C (IL-17C), also known as Cytokine CX2, is a Tcell-derived cytokine that shares sequence similarity with IL17. Thiscytokine releases tumor necrosis factor alpha and interleukin 1 betafrom a monocytic cell line. Expression of this cytokine is restricted toactivated T cells. In some instances, IL-17C has Gene ID: 27189.

Interleukin 17F (IL-17F), also known as Cytokine ML-1 or CANDF6, is acytokine that shares sequence similarity with IL17. This cytokine isexpressed by activated T cells, and stimulates the production of severalother cytokines, including IL6, IL8, and CSF2/GM_CSF. This cytokine alsoinhibits the angiogenesis of endothelial cells and induces endothelialcells to produce IL2, TGFB1/TGFB, and monocyte chemoattractantprotein-1. In some instances, IL-17F has Gene ID: 112744.

Interleukin 17 Receptor A (IL-17RA), also known as CDw217 or IL17R, is aproinflammatory cytokine secreted by activated T-lymphocytes. It is apotent inducer of the maturation of CD34-positive hematopoieticprecursors into neutrophils. The transmembrane protein encoded by thisgene (interleukin 17A receptor; IL17RA) is a ubiquitous type I membraneglycoprotein that binds with low affinity to interleukin 17A.Interleukin 17A and its receptor play a pathogenic role in manyinflammatory and autoimmune diseases such as rheumatoid arthritis. Insome instances, IL-17RA has Gene ID: 23765.

Interleukin 17 Receptor C (IL-17RC), also known as ZcytoR14, IL17Rhom,Interleukin-17 Receptor-Like Protein, or CANDF9, encodes a single-passtype I membrane protein that shares similarity with the interleukin-17receptor (IL-17RA). The protein is expressed in nonhemopoietic tissues,and binds both IL-17A and IL-17F with similar affinities. Multiplealternatively spliced transcript variants encoding different isoformshave been detected for this gene, and soluble, secreted proteins lackingtransmembrane and intracellular domains may function as extracellularantagonists to cytokine signaling. In some instances, IL-17RC has GeneID: 84818.

Interleukin 22 (IL-22), also known as Cytokine Zcyto18, IL-TIF, IL-D110,or TIFa, is a member of the IL10 family of cytokines that mediatecellular inflammatory responses. The encoded protein functions inantimicrobial defense at mucosal surfaces and in tissue repair. Thisprotein also has pro-inflammatory properties and plays a role in thepathogenesis of several intestinal diseases. In some instances, IL-22has Gene ID: 50616.

Interleukin 23 Subunit Alpha (IL-23A), also known as IL-23, SGRF, orP19, encodes a subunit of the heterodimeric cytokine interleukin 23(IL23). IL23 is composed of this protein and the p40 subunit ofinterleukin 12 (IL12B). The receptor of IL23 is formed by the beta 1subunit of IL12 (IL12RB1) and an IL23 specific subunit, IL23R. Both IL23and IL12 can activate the transcription activator STAT4, and stimulatethe production of interferon-gamma (IFNG). In contrast to IL12, whichacts mainly on naive CD4(+) T cells, IL23 preferentially acts on memoryCD4(+) T cells. In some instances, IL-23A has Gene ID: 51561.

Interleukin 24 (IL-24), also known as ST16, MDA7, IL10B, or C49A,encodes a member of the IL10 family of cytokines. It was identified as agene induced during terminal differentiation in melanoma cells. Theprotein encoded by this gene can induce apoptosis selectively in variouscancer cells. Overexpression of this gene leads to elevated expressionof several GADD family genes, which correlates with the induction ofapoptosis. The phosphorylation of mitogen-activated protein kinase 14(MAPK7/P38), and heat shock 27 kDa protein 1 (HSPB2/HSP27) are found tobe induced by this gene in melanoma cells, but not in normal immortalmelanocytes. In some instances, IL-24 has Gene ID: 11009.

Interleukin 26 (IL-26), also known as AK155, was identified by itsoverexpression specifically in herpesvirus samimiri-transformed T cells.The encoded protein is a member of the IL10 family of cytokines. It is asecreted protein and may function as a homodimer. This protein isthought to contribute to the transformed phenotype of T cells afterinfection by herpesvirus samimiri. In some instances, IL-26 has Gene ID:55801.

Interleukin 31 (IL-31), also known as IL-31, which is made principallyby activated Th2-type T cells, interacts with a heterodimeric receptorconsisting of IL31RA (MIM 609510) and OSMR (MIM 601743) that isconstitutively expressed on epithelial cells and keratinocytes. IL31 maybe involved in the promotion of allergic skin disorders and inregulating other allergic diseases, such as asthma. In some instances,IL-31 has Gene ID: 386653.

Interleukin 33 (IL-33), also known as DVS27-Related Protein, C9orf26,IL1F11, or NFEHEV, encodes a protein that is a cytokine that binds tothe IL1RL1/ST2 receptor. The encoded protein is involved in thematuration of Th2 cells and the activation of mast cells, basophils,eosinophils and natural killer cells. Several transcript variantsencoding different isoforms have been found for this gene. In someinstances, IL-33 has Gene ID: 90865.

Interleukin 5 (IL-5), also known as Eosinophil Differentiation Factor,T-Cell Replacing Factor, B-Cell Differentiation Factor I, TRF, or EDF,encodes a cytokine that acts as a growth and differentiation factor forboth B cells and eosinophils. The encoded cytokine plays a major role inthe regulation of eosinophil formation, maturation, recruitment andsurvival. The increased production of this cytokine may be related topathogenesis of eosinophil-dependent inflammatory diseases. Thiscytokine functions by binding to its receptor, which is a heterodimer,whose beta subunit is shared with the receptors for interleukine 3 (IL3)and colony stimulating factor 2 (CSF2/GM-CSF). In some instances, IL-5has Gene ID: 3567.

Interleukin 6 (IL-6), also known as B-Cell Stimulatory Factor 2, CTLDifferentiation Factor, Hybridoma Growth Factor, or IFN-βeta-2, encodesa cytokine that functions in inflammation and the maturation of B cells.In addition, the encoded protein is an endogenous pyrogen capable ofinducing fever in subjects with autoimmune diseases or infections. Theprotein is primarily produced at sites of acute and chronicinflammation, where it is secreted into the serum and induces atranscriptional inflammatory response through interleukin 6 receptor,alpha. The functioning of this gene is implicated in a wide variety ofinflammation-associated disease states, including susceptibility todiabetes mellitus and systemic juvenile rheumatoid arthritis. In someinstances, IL-6 has Gene ID: 3569.

Interleukin 8 (IL-8), also known as CXC Motif Chemokine Ligand 8, GCP-1,or NAP-1, encodes a protein that is a member of the CXC chemokine familyand is a major mediator of the inflammatory response. The encodedprotein is secreted primarily by neutrophils, where it serves as achemotactic factor by guiding the neutrophils to the site of infection.This chemokine is also a potent angiogenic factor. This gene is believedto play a role in the pathogenesis of bronchiolitis, a commonrespiratory tract disease caused by viral infection. In some instances,IL-8 has Gene ID: 3576.

Lipocalin 2 (LCN2), also known as NGAL, P25, or 24p3, encodes a proteinthat belongs to the lipocalin family. Members of this family transportsmall hydrophobic molecules such as lipids, steroid hormones andretinoids. The protein encoded by this gene is a neutrophilgelatinase-associated lipocalin and plays a role in innate immunity bylimiting bacterial growth as a result of sequestering iron-containingsiderophores. In some instances, LCN2 has Gene ID: 3934.

S100 Calcium Binding Protein A7 (S100A7), also known as PSOR1 orPsoriasin, encodes a protein that is a member of the S100 family ofproteins containing 2 EF-hand calcium-binding motifs. S100 proteins arelocalized in the cytoplasm and/or nucleus of a wide range of cells, andinvolved in the regulation of a number of cellular processes such ascell cycle progression and differentiation. The protein is overexpressedin hyperproliferative skin diseases, exhibits antimicrobial activitiesagainst bacteria and induces immunomodulatory activities. In someinstances, S100A7 has Gene ID: 6278.

5100 Calcium Binding Protein A9 (S100A9), also known as MRP-14, CAGB, orL1 AG, encodes a protein that is a member of the 5100 family of proteinscontaining 2 EF-hand calcium-binding motifs. S100 proteins are localizedin the cytoplasm and/or nucleus of a wide range of cells, and involvedin the regulation of a number of cellular processes such as cell cycleprogression and differentiation. S100 genes include at least 13 memberswhich are located as a cluster on chromosome 1q21. This antimicrobialprotein exhibits antifungal and antibacterial activity. In someinstances, S100A9 has Gene ID: 6280.

Tumor Necrosis Factor (TNF), also known as TNF-alpha, Chachectin, orDIF, encodes a multifunctional proinflammatory cytokine that belongs tothe tumor necrosis factor (TNF) superfamily. This cytokine is mainlysecreted by macrophages. It can bind to, and thus functions through itsreceptors TNFRSF1A/TNFR1 and TNFRSF1B/TNFBR. This cytokine is involvedin the regulation of a wide spectrum of biological processes includingcell proliferation, differentiation, apoptosis, lipid metabolism, andcoagulation. This cytokine has been implicated in a variety of diseases,including autoimmune diseases, insulin resistance, and cancer. In someinstances, TNF has Gene ID: 7124.

TNF Receptor Superfamily Member 1A (TNFRSF1A), also known as TNF-R1,P55, P60, or CD120a Antigen, encodes a member of the TNF receptorsuperfamily of proteins. The encoded receptor is found in membrane-boundand soluble forms that interact with membrane-bound and soluble forms,respectively, of its ligand, tumor necrosis factor alpha. Binding ofmembrane-bound tumor necrosis factor alpha to the membrane-boundreceptor induces receptor trimerization and activation, which plays arole in cell survival, apoptosis, and inflammation. Proteolyticprocessing of the encoded receptor results in release of the solubleform of the receptor, which can interact with free tumor necrosis factoralpha to inhibit inflammation. Mutations in this gene may also beassociated with multiple sclerosis in human patients. In some instances,TNFRSF1A has Gene ID: 7132.

Thymic Stromal Lymphopoietin (TSLP) encodes a hemopoietic cytokineproposed to signal through a heterodimeric receptor complex composed ofthe thymic stromal lymphopoietin receptor and the IL-7R alpha chain. Itmainly impacts myeloid cells and induces the release of Tcell-attracting chemokines from monocytes and enhances the maturation ofCD11c(+) dendritic cells. The protein promotes T helper type 2 (TH2)cell responses that are associated with immunity in various inflammatorydiseases, including asthma, allergic inflammation and chronicobstructive pulmonary disease. In some instances, TSLP has Gene ID:85480.

C-C Motif Chemokine Ligand 17 (CCL17), also known as Small-InducibleCytokine A17, TARC, or ABCD-2, is one of several Cys-Cys (CC) cytokinegenes clustered on the q arm of chromosome 16. The CC cytokines areproteins characterized by two adjacent cysteines. The cytokine encodedby this gene displays chemotactic activity for T lymphocytes, but notmonocytes or granulocytes. The product of this gene binds to chemokinereceptors CCR4 and CCR8. This chemokine plays important roles in T celldevelopment in thymus as well as in trafficking and activation of matureT cells. In some instances, TSLP has Gene ID: 85480.

C-C Motif Chemokine Ligand 18 (CCL18), also known as SCYA18, CD-CK1, orAMAC1, is one of several Cys-Cys (CC) cytokine genes clustered on the qarm of chromosome 17. The CC cytokines are proteins characterized by twoadjacent cysteines. The cytokine encoded by this gene displayschemotactic activity for naive T cells, CD4+ and CD8+ T cells andnonactivated lymphocytes, but not for monocytes or granulocytes. Thischemokine attracts naive T lymphocytes toward dendritic cells andactivated macrophages in lymph nodes. In some instances, CCL18 has GeneID: 6362.

C-C Motif Chemokine Ligand 19 (CCL19), also known as CK Beta-11,Exodus-3, or MIP3B, is one of several CC cytokine genes clustered on thep-arm of chromosome 9. The CC cytokines are proteins characterized bytwo adjacent cysteines. The cytokine encoded by this gene may play arole in normal lymphocyte recirculation and homing. It also plays animportant role in trafficking of T cells in thymus, and in T cell and Bcell migration to secondary lymphoid organs. It specifically binds tochemokine receptor CCR7. In some instances, CCL19 has Gene ID: 6363.

C-C Motif Chemokine Ligand (CCL26), also known as MacrophageInflammatory Protein 4-Alpha, Eotaxin-3, or SCYA26, is one of twoCys-Cys (CC) cytokine genes clustered on the q arm of chromosome 7. TheCC cytokines are proteins characterized by two adjacent cysteines. Thecytokine encoded by this gene displays chemotactic activity for normalperipheral blood eosinophils and basophils. The product of this gene isone of three related chemokines that specifically activate chemokinereceptor CCR3. This chemokine may contribute to the eosinophilaccumulation in atopic diseases. In some instances, CCL26 has Gene ID:10344.

C-C Motif Chemokine Ligand (CCL27), also known as Skinkine, IL-11,Ralpha-Locus Chemokine, or CTACK, is one of several CC cytokine genesclustered on the p-arm of chromosome 9. The CC cytokines are proteinscharacterized by two adjacent cysteines. The protein encoded by thisgene is chemotactic for skin-associated memory T lymphocytes. Thiscytokine may also play a role in mediating homing of lymphocytes tocutaneous sites. It specifically binds to chemokine receptor 10 (CCR10).Studies of a similar murine protein indicate that these protein-receptorinteractions have a pivotal role in T cell-mediated skin inflammation.In some instances, CCL27 has Gene ID: 10850.

C-X-C Motif Chemokine Ligand 10 (CXCL10), also known as Gamma IP10,SCYB10, or Crg-2, encodes a chemokine of the CXC subfamily and ligandfor the receptor CXCR3. Binding of this protein to CXCR3 results inpleiotropic effects, including stimulation of monocytes, natural killerand T-cell migration, and modulation of adhesion molecule expression. Insome instances, CXCL10 has Gene ID: 3627.

C-X-C Motif Chemokine Ligand 11 (CXCL11), also known as Beta-R1, SCYB11,or ITAC, is a CXC member of the chemokine superfamily. Its encodedprotein induces a chemotactic response in activated T-cells and is thedominant ligand for CXC receptor-3. IFN-gamma is a potent inducer oftranscription of this gene. In some instances, CXCL11 has Gene ID: 6373.

C-X-C Motif Chemokine Ligand 9 (CXCL9), also known as Small-InducibleCytokine B9, SCYB9, Crg-10, or HuMIG, encodes a protein thought to beinvolved in T cell trafficking. The encoded protein binds to C-X-C motifchemokine 3 and is a chemoattractant for lymphocytes but not forneutrophils. In some instances, CXCL9 has Gene ID: 4283.

Interleukin 13 (IL-13) encodes an immunoregulatory cytokine producedprimarily by activated Th2 cells. This cytokine is involved in severalstages of B-cell maturation and differentiation. It up-regulates CD23and MHC class II expression, and promotes IgE isotype switching of Bcells. This cytokine down-regulates macrophage activity, therebyinhibits the production of pro-inflammatory cytokines and chemokines.This cytokine is found to be critical to the pathogenesis ofallergen-induced asthma but operates through mechanisms independent ofIgE and eosinophils. This gene, IL3, IL5, IL4, and CSF2 form a cytokinegene cluster on chromosome 5q, with this gene particularly close to IL4.In some instances, IL-13 has Gene ID: 3596.

Interleukin 13 Receptor (IL-13R) is a type I cytokine receptor, bindingInterleukin-13. It consists of two subunits, encoded by IL13RA1 andIL4R, respectively. These two genes encode the proteins IL-13Rα1 andIL-4Rα. These form a dimer with IL-13 binding to the IL-13Rα1 chain andIL-4Rα stabilises this interaction.

Interleukin 4 (IL-4), also known as Lymphocyte Stimulatory Factor 1,BCGF-1, or BSF1, encodes a protein that is a pleiotropic cytokineproduced by activated T cells. This cytokine is a ligand for interleukin4 receptor. The interleukin 4 receptor also binds to IL13, which maycontribute to many overlapping functions of this cytokine and IL13.STAT6, a signal transducer and activator of transcription, has beenshown to play a central role in mediating the immune regulatory signalof this cytokine. This gene, IL3, IL5, IL13, and CSF2 form a cytokinegene cluster on chromosome 5q, with this gene particularly close toIL13. This gene, IL13 and IL5 are found to be regulated coordinately byseveral long-range regulatory elements in an over 120 kilobase range onthe chromosome. In some instances, IL-4 has Gene ID: 3565.

Nitric Oxide Synthetase 2 (NOS2), also known as Inducible NOS2 orHepatocyte NOS, encodes a nitric oxide synthase which is expressed inliver and is inducible by a combination of lipopolysaccharide andcertain cytokines. In some instances, NOS2 has Gene ID: 4843.

5100 Calcium Binding Protein A8 (S100A8), also known as Leukocyte L1Complex Light Chain, Cystic Fibrosis Antigen, or Calgranulin A, encodesa protein that is a member of the S100 family of proteins containing 2EF-hand calcium-binding motifs. This protein may function in theinhibition of casein kinase and as a cytokine. Altered expression ofthis protein is associated with the disease cystic fibrosis. In someinstances, S100A8 has Gene ID: 6279.

Adenosine Deaminase, RNA Specific (ADAR), also known asInterferon-Inducible Protein 4, K88DSRBP, or P136, encodes the enzymeresponsible for RNA editing by site-specific deamination of adenosines.This enzyme destabilizes double-stranded RNA through conversion ofadenosine to inosine. Mutations in this gene have been associated withdyschromatosis symmetrica hereditaria. In some instances, ADAR has GeneID: 103.

Autophagy Related 5 (ATG5), also known as Apoptosis-Specific Protein,APG51, or HAPG5, encodes a protein that, in combination with autophagyprotein 12, functions as an E1-like activating enzyme in aubiquitin-like conjugating system. The encoded protein is involved inseveral cellular processes, including autophagic vesicle formation,mitochondrial quality control after oxidative damage, negativeregulation of the innate antiviral immune response, lymphocytedevelopment and proliferation, MHC II antigen presentation, adipocytedifferentiation, and apoptosis. In some instances, ATG5 has Gene ID:9474.

C-C Motif Chemokine Ligand 5 (CCL5), also known as SIS-Delta, EosinophilCheomtactic Cytokine, or TCP228, is one of several chemokine genesclustered on the q-arm of chromosome 17. This chemokine, a member of theCC subfamily, functions as a chemoattractant for blood monocytes, memoryT helper cells and eosinophils. It causes the release of histamine frombasophils and activates eosinophils. This cytokine is one of the majorHIV-suppressive factors produced by CD8+ cells. It functions as one ofthe natural ligands for the chemokine receptor chemokine (C-C motif)receptor 5 (CCR5), and it suppresses in vitro replication of the R5strains of HIV-1, which use CCR5 as a coreceptor. In some instances,CCL5 has Gene ID: 6352.

The interferon-inducible p200 family of proteins (IFI's) are geneproducts induced by interferons (IFNs). Proteins in this family sharesignificant homology, with human homologues comprising IFI-16, myeloidcell nuclear differentiation antigen (MNDA) and absent in melanoma (AIM)2. The p200 proteins have been implicated in cell cycle regulation anddifferentiation based on their ability to interact with and modulate theactivities of multiple transcriptional factors such as Rb and p53

Interferon Induced proteins with Tetratricopeptide repeats (IFIT's)confer immunity against viral infection. These proteins are generallyproduced during viral infection, Interferon (IFN) treatment, and duringpathogen recognition (Pathogen associated molecular pattern recognition)by the immune system during infections.

Interferon Alpha 1 (IFNA1), also known as Interferon Alpha-D, IFNA13, orLelFD, encodes a protein that is produced by macrophages and hasantiviral activity. In some instances, IFNA1 has Gene ID: 3439.

Interferon Alpha 2 (IFNA2), also known as Alpha-2a Interferon, IFN2B, orIFNA2C, is a member of the alpha interferon gene cluster on chromosome9. The encoded protein is a cytokine produced in response to viralinfection. Use of the recombinant form of this protein has been shown tobe effective in reducing the symptoms and duration of the common cold.In some instances, IFNA2 has Gene ID: 3440.

Interferon Alpha 4 (IFNA4), also known as Interferon Alpha-4B,Interferon Alpha-M1 or Interferon Alpha-76, is a Protein Coding gene.Diseases associated with IFNA4 include Rabies. Among its relatedpathways are RIG-I/MDA5 mediated induction of IFN-alpha/beta pathwaysand Cytokine Signaling in Immune system. In some instances, IFNA4 hasGene ID: 3441.

Interferon Alpha And Beta Receptor Subunit 1 (IFNAR1), also known asCytokine Receptor Class-II Member 1, IFN-R-1, or AVP, encodes a proteinthat is a type I membrane protein that forms one of the two chains of areceptor for interferons alpha and beta. Binding and activation of thereceptor stimulates Janus protein kinases, which in turn phosphorylateseveral proteins, including STAT1 and STAT2. The encoded protein alsofunctions as an antiviral factor. In some instances, IFNAR1 has Gene ID:3454.

Interferon Alpha And Beta Receptor Subunit (IFNAR2), also known asIFNABR, Interferon Alpha Binding Protein, or IMD45, encodes a proteinthat is a type I membrane protein that forms one of the two chains of areceptor for interferons alpha and beta. Binding and activation of thereceptor stimulates Janus protein kinases, which in turn phosphorylateseveral proteins, including STAT1 and STAT2. In some instances, IFNAR2has Gene ID: 3455.

Interferon Beta 1 (IFNB1), also known as Fibroblast Interferon,IFN-Beta, or IFF, encodes a cytokine that belongs to the interferonfamily of signaling proteins, which are released as part of the innateimmune response to pathogens. The protein encoded by this gene areinvolved in cell differentiation and anti-tumor defenses. Followingsecretion in response to a pathogen, type I interferons bind ahomologous receptor complex and induce transcription of genes such asthose encoding inflammatory cytokines and chemokines. Overactivation oftype I interferon secretion is linked to autoimmune diseases. In someinstances, IFNB1 has Gene ID: 3456.

Interferon Epsilon (IFNE), also known as Interferon Tau-1 or PRO655, isa Protein Coding gene. Among its related pathways are PEDF InducedSignaling and JAK-STAT signaling pathway (KEGG). In some instances, IFNEhas Gene ID: 338376.

Interferon Omega 1 (IFNW1), also known as Interferon Alpha-II-1, encodesa protein that is an interferon and possesses antiviral activity. Theencoded protein binds to the interferon alpha/beta receptor but not tothe interferon gamma receptor. In some instances, IFNW1 has Gene ID:3467.

Interleukin 1 Receptor Associated Kinase 1 (IRAK1), also known asEC2.7.11.1 or Pelle, encodes the interleukin-1 receptor-associatedkinase 1, one of two putative serine/threonine kinases that becomeassociated with the interleukin-1 receptor (IL1R) upon stimulation. Thisgene is partially responsible for IL1-induced upregulation of thetranscription factor NF-kappa B. In some instances, IRAK1 has Gene ID:3654.

Interferon Regulatory Factors (IRF's) are proteins which regulatetranscription of interferons. They are used in the JAK-STAT signalingpathway. Expression of IRF genes is under epigenetic regulation bypromoter DNA methylation.

Keratins (KRT's) are a family of fibrous structural proteins making uphair, nails, horns, claws, hooves, and the outer layer of human skin.Keratin is also the protein that protects epithelial cells from damageor stress.

2′-5′-Oligoadenylate Synthetase 1 (OAS1), also known as E18/E16, OIAS,or P46/P42 OAS, is induced by interferons and encodes a protein thatsynthesizes 2′,5′-oligoadenylates (2-5As). This protein activates latentRNase L, which results in viral RNA degradation and the inhibition ofviral replication.

Osteopontin (OPN), also known as bone sialoprotein I (BSP-1 or BNSP),early T-lymphocyte activation (ETA-1), secreted phosphoprotein 1 (SPP1),2ar and Rickettsia resistance (Ric), is a protein that in humans isencoded by the SPP1 gene (secreted phosphoprotein 1). In some instances,SPP1 has Gene ID: 6696.

Signal Transducer And Activator Of Transcription 4 (STAT4), also knownas SLEB11, encodes a protein that is a member of the STAT family oftranscription factors. In response to cytokines and growth factors, STATfamily members are phosphorylated by the receptor associated kinases,and then form homo- or heterodimers that translocate to the cell nucleuswhere they act as transcription activators. This protein is essentialfor mediating responses to IL12 in lymphocytes, and regulating thedifferentiation of T helper cells. Mutations in this gene may beassociated with systemic lupus erythematosus and rheumatoid arthritis.In some instances, STAT4 has Gene ID: 6775.

TNF Alpha Induced Protein 3 (TNFAIP3), also known as PutativeDNA-Binding Protein A20, Zinc Finger Protein A20, or OTUD7C, is a genewhose expression is rapidly induced by the tumor necrosis factor (TNF).The protein encoded by this gene is a zinc finger protein andubiqitin-editing enzyme, and has been shown to inhibit NF-kappa Bactivation as well as TNF-mediated apoptosis. The encoded protein, whichhas both ubiquitin ligase and deubiquitinase activities, is involved inthe cytokine-mediated immune and inflammatory responses. In someinstances, TNFAIP3 has Gene ID: 7128.

Tyrosine Kinase 2 (TYK2), also known as Non-Receptor Tyrosine-ProteinKinase TYK2, EC 2.7.10.2, or IMD35, encodes a member of the tyrosinekinase and, more specifically, the Janus kinases (JAKs) proteinfamilies. This protein associates with the cytoplasmic domain of type Iand type II cytokine receptors and promulgate cytokine signals byphosphorylating receptor subunits. It is also component of both the typeI and type III interferon signaling pathways. As such, it may play arole in anti-viral immunity. In some instances, TYK2 has Gene ID: 7297.

EXAMPLES

These examples are provided for illustrative purposes only and not tolimit the scope of the claims provided herein.

Example 1—Non-Invasive Gene Expression Analysis for Psoriasis

Progress has been made in the treatment of moderate to severe psoriasisby blocking TNF alpha, IL-17A and IL-23. The pathways affected aredepicted in FIG. 6. Further, disease monitoring, prediction of flare-upsand treatment selection remain challenging. Described below is anon-invasive method to assess gene expression in psoriasis and topredict treatment response.

Samples were assayed using the adhesive patch-based skin biopsy platformdescribed herein. The modular structure of the qRT-PCR assay allowed itto be employed in a number of inflammatory skin conditions includingpsoriasis, atopic dermatitis, or lupus. In psoriasis, the assay focusedon 20 targets involved in expanded TH17 pathways.

Over 500 lesional and non-lesional adhesive patch biopsy samples frompatients with moderate to severe psoriasis demonstrated detection of 20selected targets by qRT-PCR and differences in gene expressionsignatures of lesional, non-lesional, and non-psoriasis control skin(p<0.001, n=24). Analyses from non-lesional samples avoided the need tocontrol for disease activity in individual psoriasis lesions and provideclinically useful information. In non-lesional psoriatic sample comparedto normal skin, gene expression levels of IL-17A, IL-17C, IL-17F, IL-17receptors, IL-23A, IL-22, IL-24, IL-6, IL-8, CXCL1, CXCL5, DEFB4A, LCN2,S100A7 as well as TNF-α and its receptor were altered by 2 to 200 fold.Therapeutic intervention with targeted therapeutics such as ixekizumab(p<0.001) reduced target gene expression (including IL-17A and 17F,TNF-α and CXCL1 and CXCL5) compared to baseline after 2 weeks.

Results indicate non-invasive gene expression analysis of lesional andnon-lesional epidermal skin samples is a method to monitor diseaseactivity with the potential to predict flare-ups and treatment failurein psoriasis.

Example 2—Non-Invasive Gene Expression Analysis for Psoriasis Utilizinga Different Test Population

An adhesive patch-based device as described herein was used to collectepidermal skin cells from test subjects (healthy persons as control andtreatment naïve psoriatic patients) through a non-invasive procedure(from psoriatic patients, both lesional and non-lesional skins were alsocollected). Total RNA was extracted from these samples and used forcytokine gene expression analysis with TaqMan RT-qPCR. A panel of 13cytokines, mainly in the Th17 pathway, involved in psoriasis was studiedand their expression levels were calculated through the threshold cyclecount (Ct) from the qPCR.

With the adhesive patch-based device, epidermal tissues were collectedfrom all test subjects and total RNA was isolated (FIGS. 28A-28C). Aspsoriatic lesion often had a thickened skin with dried flaky layers oftissues, adhesive patch sampling often yielded more skin tissues thushigher RNA yields from psoriatic lesional skins than that from normal ornon-lesional skins.

Testing of isolated RNA by qPCR allowed for detection of gene expressionchanges. FIGS. 29A-29F show tests on several key genes (IL-17, IL-23,DEFB4 and S100A9) and compares their expression levels in both PSOR andNS (normal skin) with dilutions of RNA inputs in qPCR. Elevated geneexpressions (shown as a downward shift of Ct value) are seen in PSORskins for most targets, while a linear parallel changes in Ct in bothtarget gene and housekeeping gene ACTB with changing RNA input in qPCRconfirms both the quality of the isolated RNA for gene expression andaccuracy of the qPCR analysis adopted to the current assay on cytokinegene expression analysis.

Psoriasis is affected by many cytokines and their interactions. FIG. 30shows a heatmap constructed from the Ct values of 13 cytokine genes from53 RNA samples (14 NML, 15 NL and 24 PSOR skins). A darker red on theheatmap shows a lower Ct or an increased gene expression while a darkergrey shows a higher Ct or a lower gene expression in the cells. Thepsoriatic lesional skins have demonstrated a different heatmap from thatin other 2 types of skins (normal and non-lesion). The gene expressionpattern within PSRO group had also displayed a high degree ofuniformity, in comparison with traditional biopsy method such as liquidbiopsy or surgical biopsy.

In addition, the non-invasive gene expression analysis assay had alsodetected subgroups of PSOR lesions, varied in the expression levels ofkey genes (IL-17, 22, 26) in the Th17 pathway (FIG. 31), which mightassociate to patients who fail to respond to drug treatment.

Example 3—Stratum Corneum Gene Expression Measurements in PsoriasisLesional and Non-Lesional Skin

Samples were collected using the adhesive patch-based device describedherein. Antibodies selective against cytokine targets including theirreceptors were assessed for expression levels in 24 psoriasis lesionaland 15 non-lesional skin samples. Samples were tested for binding toIL-17RC, IL-26, IL-22, IL-17A, IL-17F, IL-17C, TNFα, IL-6, IL-23A,DEFB4A, S100A9, CXCL5, and IL-8. FIG. 5 is a heat map showing results ofscreening. In psoriatic samples, lower levels of antibody binding weredetected to DEFB4A, S100A9, CXCL5, and IL-8. In non-psoriatic samples,increased binding to IL-17RC, IL-26, IL-22, IL-17A, IL-17F, IL-17C,TNFα, IL-6, and IL-23A was detected.

Example 4—Cytokine Transcription Levels from Adhesive Patch Samples

Two RNA samples were collected from psoriatic skin using adhesive patchcollection methods described herein. Table 1 shows elevated expressionlevels of psoriatic cytokines from the IL-23/TH17 axis.

TABLE 1 IL- Sample 17A 17RA IL-17C 17RC IL-17F TNF-a S100A7 S100A9 1ΔΔCt −3.7 0.6 -6.3 1.0 −2.7 −8.3 −12.9 −17.5 Fc 12.6 0.7 76.7 0.5 6.6314.7 7781.4 186063.8 2 ΔΔCt −7.4 −0.8 −6.6 1.2 −2.9 −6.3 −14.1 −16.7 FC168.9 1.7 97.2 0.4 7.2 81.2 17104.6 109218.3 Sample CCL20 IL-22 CXCL1IL-24 CXCL5 IL-26 LCN2 DEFB4A 1 ΔΔCt −1.9 −7.7 −7.0 −0.4 −7.9 −2.5 −10.2−17.4 Fc 3.7 208.5 128.6 1.3 239.0 5.5 1171.8 169438.3 2 ΔΔCt 6.0 0.2−3.1 0.6 −9.7 −6.9 −10.2 −18.5 FC 0.0 0.9 8.8 0.7 827.0 122.7 1172.4368594.1 Note: ΔΔCt = (ΔCt_lesion-ΔCt_normal); FC (fold of change) =2{circumflex over ( )} ΔΔCt

Example 5—Gene Expression Changes in Psoriasis

Samples were collected and assayed according to the methods describedherein. The fold change of gene expression level in psoriatic lesionskin compared to normal skin and in non-lesional skin compared to normalskin were calculated. FIG. 25 shows the cytokines with increase geneexpression detected in both lesional skin and non-lesion area. FIG. 26shows cytokines with decreased gene expression in uninvolvednon-lesional skin but increased gene expression in psoriatic lesionalskin.

Example 6—Expanded TH2 Assay for Atopic Dermatitis

Samples were collected using the adhesive patch-based skin biopsyplatform described herein and assayed. The modular structure of theqRT-PCR assay allows it to be employed in a number of inflammatory skinconditions including psoriasis, atopic dermatitis or lupus. In atopicdermatitis, the assay focused on 18 targets involved in expanded TH2pathways (see FIG. 7). Targets included IL-4, IL-13, IL-17, IL-22,IL-31, TSLP, CXCL9, CXCL10, CXCL11, S100A7, S100A8, S100A9, CCL17,CCL18, CCL19, CCL26, CCL27, and NOS2.

Samples were screened from 39 subjects with atopic dermatitis. As shownin FIG. 11, IL-13 expression was detected in less than half (about 41%)of the subjects. No samples showed IL-4 expression. 100% of samplesshowed IL-13/4 receptor expression and about 95% exhibited CCL17expression.

Results suggest selecting for patients with IL-13 expression will leadto a higher proportion of responders to treatment with receptor blockingagents.

Example 7—Expression Levels in AD Samples

Samples from AD subjects were collected and assayed according to themethods described herein. Forty samples from lesional areas, 17 samplesfrom non-lesional areas, and 20 samples from normal skin were assayedfor expression levels of IL-31RA, CCL17, IL-23A, IL-4R, IL22, IL-13, andIL-13RA1, see FIGS. 27A-27I.

Example 8—IL-13/4 Receptor Blocking with Dupilumab

AD subjects were treated with 300 mg dupilumab with subcutaneousadministration every other week for 16 weeks. Samples were collected asdescribed in Example 5.

About 50% of test subjects achieved 75% reduction in symptoms (EASI-75)compared to about 15% of placebo subjects achieving EASI-75 (FIG. 8).Additionally, about 38% of subjects achieved an IGA score of cleared (0)or minimal (1) (IGA 0-1) compared to about 10% of placebo subjects.

Example 9—IL-13 Blocking with Lebrikizumab

AD subjects were treated with 125 mg lebrikizumab, an IL-13 blockingmonoclonal antibody, and corticosteroids weekly for 12 weeks. Sampleswere collected as described in Example 5.

About 55% of test subjects achieved EASI-75 compared to about 34% ofplacebo subjects (n-50/group, p=0.05) (see FIG. 9)

Example 10—IL-13 Blocking with Tralokinumab

AD subjects were tested for DPP-4 levels in the blood. Treatment andplacebo groups were further selected for elevated DPP-4 levels.Treatment groups received tralokinumab, an IL-13 blocking monoclonalantibody, and topical steroids for 12 weeks. Samples were collected asdescribed in Example 5.

As shown in FIG. 10, groups not selected for elevated DPP-4 levels,about 26% of treatment subjects achieved IGA 0-1 compared to about 12%of placebo subjects. In groups selected for elevated DPP-4 levels, about35% of test subjects achieved IGA 0-1 compared to IGA 0-1 8% of placebosubjects. In groups selected for elevated DPP-4 levels, about 52% oftreatment subjects achieved EASI-75 as compared to about 13% of placebosubjects.

Results show tralokinumab resolves AD symptoms in subject and resolvessymptoms to a higher degree in subjects exhibiting elevated DPP-4 levelsin the blood.

Example 11—Expression of CCL17 in AD Lesion and Non-Lesion Areas

AD samples were collected from using the adhesive patch-based skinbiopsy platform described herein and assayed. Lesional and non-lesionalareas as well as normal skin were tested for CCL17 expression (FIG. 12Aand FIG. 12B). ΔCt analysis shows that although changes are more severein lesional skin samples, the changing patterns are similar in both thelesional and non-lesional samples from AD subjects, suggestingnon-lesional samples may be used as a mode of disease diagnosis.ΔCt=normalized gene expression change (=CT_(target gene)−Ct_(ACTB)). Alarger ΔCt value means less expression of the target gene. A smaller ΔCtvalue means more expression of the target gene.

Example 12—Expression Levels of IL-13, IL-22, and IL-23A in AD Samples

AD samples were collected from using the adhesive patch-based skinbiopsy platform described herein and assayed. Lesional and non-lesionalareas as well as normal skin were tested for IL-13, IL-22, and IL-23Aexpression levels (FIGS. 13A, 13B, 14A, 14B, 15A, and 15B). ΔCt analysisshows all three cytokines have very low gene expression levels inhealthy skin samples (triangles), but show different gene expressionpatterns in different AD samples (diamonds). Expression levels wereeither remarkably increased, as indicated by a reduced Ct, or remainedunchanged, as shown by a high Ct. It is considered that differentialgene expression may be related to “responders” or “non-responders” ofthe disease and as such, IL-13, IL-22, and IL-23A expression is apotential for screening responders from non-responders.

Example 13—Expression Levels of IL-31 and IL-31R in AD Samples

AD samples were collected from using the adhesive patch-based skinbiopsy platform described herein and assayed. Lesional and non-lesionalareas as well as normal skin were tested for IL-31 and IL-31R expressionlevels (FIGS. 16A, 16B, 17A, 17B, 18A, and 18B). Similar to cytokineIL-4, IL-31 is another cytokine important in regulating AD disease. Someincrease in IL-31 is detected in lesional samples. IL-31 receptor(IL-31R) shows reduced gene expression in lesional samples.

Example 14—Expression Levels of IL-13 and IL31RA1 in AD Samples

IL-13 and IL-4 are proposed to work in AD according to the pathwaydepicted in FIG. 19. AD samples were collected from using the adhesivepatch-based skin biopsy platform described herein and assayed. Lesionaland non-lesional areas as well as normal skin were tested for IL-13,IL-13RA1, and IL4R expression. Results are shown in FIGS. 20A, 20B, 21A,21B, 22A, and 22B. An increase in IL-13 expression was detected andaccompanied by a decrease in the gene expression of its receptor,IL-13RA1. IL-4 gene expression was not detected in the samples, whileIL-4R showed expression level that remained unchained in AD samples ascompared to normal skin.

These results suggest that IL-13 plays a more significant role that IL-4in AD disease regulation.

Example 15—Expression Levels of NOS2 in AD Samples

AD samples were collected from using the adhesive patch-based skinbiopsy platform described herein and assayed for NOS2 expression levels.Results in FIG. 23 show AD samples with a lower average Ct compared tonormal skin samples, indicating an increased expression level in ADsamples.

Example 16—Expanded Interferon Response Gene Assay for Lupus

Samples are collected using the adhesive patch-based skin biopsyplatform described herein and assayed. The modular structure of theqRT-PCR assay allows it to be employed in a number of inflammatory skinconditions including psoriasis, atopic dermatitis or lupus. In lupus,the assay focused on 21 targets involved in a neutrophil-mediated flare(see FIG. 24). Targets include IFNA1, IFNA2, IFNA4, IFNAR1, IFNAR2,IFNB1, IFNE, IFNW1, ADAR, CCL5, IFIT's, IFI's, IRF's, OAS1, IRAK1,TNFAIP3, ATG5, TYK2, STAT4, OPN, and KRT's.

Lesional and non-lesional adhesive patch biopsy samples from patientswith moderate to severe lupus are tested against 21 selected targets byqRT-PCR to show differences in gene expression signatures of lesional,non-lesional and non-lupus control skin.

Embodiment 1

A method of detecting gene expression levels of at least two of IL-17A,IL-17F, IL-8, CXCL5, S100A9, and DEFB4A in a subject suspected of havingpsoriasis, comprising: (a) isolating nucleic acids from a skin sampleobtained from the subject, where the skin sample comprises cells fromthe stratum corneum; and (b) detecting the expression levels of at leasttwo of IL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4A by contacting theisolated nucleic acids with a set of probes that recognizes at least twoof IL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4A, and detect bindingbetween at least two of IL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4Aand the set of probes.

Embodiment 2

The method of embodiment 1, wherein the method comprises detecting theexpression levels of at least three, at least four, or at least five ofIL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4A.

Embodiment 3

The method of embodiment 1, wherein the method comprises detecting theexpression levels of IL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4A.

Embodiment 4

The method of embodiment 1, wherein the method comprises detecting theexpression levels of IL-17A, IL-17F, IL-8, CXCL5, and S100A9.

Embodiment 5

The method of embodiment 1, wherein the method comprises detecting theexpression levels of IL-17A, IL-17F, IL-8, and CXCL5.

Embodiment 6

The method of embodiment 1, wherein the method comprises detecting theexpression levels of IL-17A, IL-17F, and IL-8.

Embodiment 7

The method of embodiment 1, wherein the method comprises detecting theexpression levels of IL-17A, and IL-17F.

Embodiment 8

The method of any one of the embodiments 1-7, wherein the expressionlevel is an up-regulated gene expression level, compared to a geneexpression level of an equivalent gene from a control sample.

Embodiment 9

The method of embodiment 8, wherein the gene expression levels ofIL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4A are upregulated.

Embodiment 10

The method of any one of the embodiments 1-9, wherein the set of probesrecognizes at least two but no more than six genes.

Embodiment 11

The method of embodiment 1, wherein the detecting comprises contactingthe isolated nucleic acids with an additional set of probes thatrecognizes IL-17C, S100A7, IL-17RA, IL-17RC, IL-23A, IL-22, IL-26,IL-24, IL-6, CXCL1, TNFα, LCN2, CCL20, TNFRSF1A, or a combinationthereof.

Embodiment 12

The method of embodiment 11, wherein the additional set of probesrecognizes one but no more than fourteen genes.

Embodiment 13

A method of detecting gene expression levels from a first geneclassifier and a second gene classifier in a subject suspected of havingpsoriasis, comprising: (a) isolating nucleic acids from a skin sampleobtained from the subject, wherein the skin sample comprises cells fromthe stratum corneum; (b) detecting the expression levels of one or moregenes from the first gene classifier: IL-17A, IL-17F, IL-8, CXCL5,S100A9, and DEFB4A, by contacting the isolated nucleic acids with a setof probes that recognizes one or more genes from the first geneclassifier, and detects binding between one or more genes from the firstgene classifier and the set of probes; and (c) detecting the expressionlevels of one or more genes from the second gene classifier: IL-17C,S100A7, IL-17RA, IL-17RC, IL-23A, IL-22, IL-26, IL-24, IL-6, CXCL1,IFN-gamma, IL-31, IL-33, TNFα, LCN2, CCL20, and TNFRSF1A, by contactingthe isolated nucleic acids with an additional set of probes thatrecognizes one or more genes from the second gene classifier, anddetects binding between one or more genes from the second geneclassifier and the additional set of probes.

Embodiment 14

The method of embodiment 13, wherein the method comprises detecting theexpression levels of IL-17A and IL-17F from the first gene classifier.

Embodiment 15

The method of embodiment 13, wherein the method comprises detecting theexpression levels of IL-8, CXCL5, S100A9, and DEFB4A from the first geneclassifier.

Embodiment 16

The method of embodiment 13, wherein the method comprises detecting theexpression levels of IL-17A, IL-8, and DEFB4A from the first geneclassifier.

Embodiment 17

The method of embodiment 13, wherein the method comprises detecting theexpression levels of IL-17F, CXCL5, and S100A9 from the first geneclassifier.

Embodiment 18

The method of embodiment 13, wherein the method comprises detecting theexpression levels of IL-17A, IL-17F, IL-8, CXCL5, S100A9, and DEFB4Afrom the first gene classifier.

Embodiment 19

The method of any one of the embodiments 13-18, wherein the expressionlevel is an up-regulated gene expression level, compared to a geneexpression level of an equivalent gene from a control sample.

Embodiment 20

The method of embodiment 19, wherein the gene expression level ofIL-17A, IL-17F, IL-8, CXCL5, S100A9, or DEFB4A is up-regulated.

Embodiment 21

The method of any one of the embodiments 13-20, wherein the set ofprobes recognizes at least one but no more than six genes.

Embodiment 22

The method of any one of the embodiments 13-20, wherein the additionalset of probes recognizes at least one but no more than 17 genes.

Embodiment 23

The method of any one of the embodiments 13-22, wherein the methodfurther comprises determining the expression level of one or more genesfrom the second classifier are upregulated.

Embodiment 24

The method of any one of the embodiments 1-23, further comprisingadministering to the subject an inhibitor of TNFα, IL-17A, or IL-23.

Embodiment 25

The method of embodiment 24, wherein if the subject has an altered geneexpression level of at least two of IL-17A, IL-17F, IL-8, CXCL5, S100A9,and DEFB4A, the subject is administered with an inhibitor of TNFα, orthe level of the treatment is increased.

Embodiment 26

The method of embodiment 24, wherein if the subject has an altered geneexpression level of at least two of IL-17A, IL-17F, IL-8, CXCL5, S100A9,and DEFB4A, the subject is administered with an inhibitor of IL-17A, orthe level of the treatment is increased.

Embodiment 27

The method of embodiment 24, wherein if the subject has an altered geneexpression level of at least two of IL-17A, IL-17F, IL-8, CXCL5, S100A9,and DEFB4A, the subject is administered with an inhibitor of IL-23, orthe level of the treatment is increased.

Embodiment 28

A method of treating a subject with an inhibitor of TNFα, IL-17A, orIL-23, wherein the subject has psoriasis, the method comprising thesteps of:

determining whether the subject has an altered gene expression level by:

isolating nucleic acids from a skin sample comprising cells from thestratum corneum; and

performing or having performed an expression analysis on the skin sampleby contacting the isolated nucleic acids with a set of probes thatrecognizes at least two of IL-17A, IL-17F, IL-8, CXCL5, S100A9, andDEFB4A, and detect binding between at least two of IL-17A, IL-17F, IL-8,CXCL5, S100A9, and DEFB4A and the set of probes; and if the subject hasan altered gene expression level of at least two of IL-17A, IL-17F,IL-8, CXCL5, S100A9, and DEFB4A, then administer to the subject aninhibitor of TNFα, IL-17A, or IL-23 or increase the level of thetreatment with the inhibitor, and if the subject does not have analtered gene expression level of at least two of IL-17A, IL-17F, IL-8,CXCL5, S100A9, and DEFB4A, then does not administer the inhibitor ordiscontinue the treatment with the inhibitor.

Embodiment 29

The method of embodiment 28, wherein if the subject has an altered geneexpression level of at least two of IL-17A, IL-17F, IL-8, CXCL5, S100A9,and DEFB4A, the subject is administered with an inhibitor of TNFα, orthe level of the treatment is increased.

Embodiment 30

The method of embodiment 29, wherein the altered gene expression is anincrease in expression.

Embodiment 31

The method of embodiment 28, wherein if the subject has an altered geneexpression level of at least two of IL-17A, IL-17F, IL-8, CXCL5, S100A9,and DEFB4A, the subject is administered with an inhibitor of IL-17A, orthe level of the treatment is increased.

Embodiment 32

The method of embodiment 31, wherein the altered gene expression is anincrease in expression.

Embodiment 33

The method of embodiment 28, wherein if the subject has an altered geneexpression level of at least two of IL-17A, IL-17F, IL-8, CXCL5, S100A9,and DEFB4A, the subject is administered with an inhibitor of IL-23, orthe level of the treatment is increased.

Embodiment 34

The method of embodiment 33, wherein the altered gene expression is anincrease in expression.

Embodiment 35

The method of embodiment 28, wherein the set of probes recognizes atleast two of IL-17A, IL-17F, and IL-8.

Embodiment 36

The method of embodiment 28, wherein the set of probes recognizes CXCL5,S100A9, and DEFB4A.

Embodiment 37

A method of detecting gene expression levels of at least two of IL-13,IL-31, and TSLP in a subject suspected of having atopic dermatitis,comprising: (a) isolating nucleic acids from a skin sample obtained fromthe subject, where the skin sample comprises cells from the stratumcorneum; and (b) detecting the expression levels of at least two ofIL-13, IL-31, and TSLP by contacting the isolated nucleic acids with aset of probes that recognizes at least two of IL-13, IL-31, and TSLP,and detect binding between at least two of IL-13, IL-31, and TSLP andthe set of probes.

Embodiment 38

The method of embodiment 37, wherein the method comprises detecting theexpression levels of at least two or at least three of IL-13, IL-31, andTSLP.

Embodiment 39

The method of embodiment 37, wherein the method comprises detecting theexpression levels of IL-13, IL-31, and TSLP.

Embodiment 40

The method of embodiment 37, wherein the method comprises detecting theexpression levels of IL-13 and IL-31.

Embodiment 41

The method of embodiment 37, wherein the method comprises detecting theexpression levels of IL-13 and TSLP.

Embodiment 42

The method of embodiment 37, wherein the expression level is anup-regulated gene expression level, compared to a gene expression levelof an equivalent gene from a control sample.

Embodiment 43

The method of embodiment 42, wherein the gene expression levels ofIL-13, IL-31, and TSLP are unregulated.

Embodiment 44

The method of embodiment 37, wherein the set of probes recognizes atleast two but no more than three genes.

Embodiment 45

The method of embodiment 37, wherein the detecting comprises contactingthe isolated nucleic acids with an additional set of probes thatrecognizes IL-13R, IL-4R, IL-17, IL-22, CXCL9, CXCL10, CXCL11, S100A7,S100A8, S100A9, CCL17, CCL18, CCL19, CCL26, CCL27, NOS2, or acombination thereof.

Embodiment 46

The method of embodiment 45, wherein the additional set of probesrecognizes one but no more than sixteen genes.

Embodiment 47

A method of detecting gene expression levels from a first geneclassifier and a second gene classifier in a subject suspected of havingatopic dermatitis, comprising: (a) isolating nucleic acids from a skinsample obtained from the subject, wherein the skin sample comprisescells from the stratum corneum; (b) detecting the expression levels ofone or more genes from the first gene classifier: IL-13, IL-31, andTSLP, by contacting the isolated nucleic acids with a set of probes thatrecognizes one or more genes from the first gene classifier, and detectsbinding between one or more genes from the first gene classifier and theset of probes; and (c) detecting the expression levels of one or moregenes from the second gene classifier: IL-13R, IL-4R, IL-17, IL-22,CXCL9, CXCL10, CXCL11, S100A7, S100A8, S100A9, CCL17, CCL18, CCL19,CCL26, CCL27, and NOS2, by contacting the isolated nucleic acids with anadditional set of probes that recognizes one or more genes from thesecond gene classifier, and detects binding between one or more genesfrom the second gene classifier and the additional set of probes.

Embodiment 48

The method of embodiment 47, wherein the method comprises detecting theexpression levels of IL-13 and IL-31 from the first gene classifier.

Embodiment 49

The method of embodiment 47, wherein the method comprises detecting theexpression levels of IL-31 and TSLP from the first gene classifier.

Embodiment 50

The method of embodiment 47, wherein the method comprises detecting theexpression levels of IL-13 and TSLP from the first gene classifier.

Embodiment 51

The method of any one of the embodiments 47-50, wherein the expressionlevel is an up-regulated gene expression level, compared to a geneexpression level of an equivalent gene from a control sample.

Embodiment 52

The method of embodiment 51, wherein the gene expression level of IL-13,IL-31, or TSLP is up-regulated.

Embodiment 53

The method of any one of the embodiments 47-52, wherein the set ofprobes recognizes at least one but no more than three genes.

Embodiment 54

The method of any one of embodiments 47-52, wherein the additional setof probes recognizes at least one but no more than 16 genes.

Embodiment 55

The method of any one of the embodiments 47-54, wherein the methodfurther comprises determining the expression level of one or more genesfrom the second classifier are unregulated.

Embodiment 56

The method of any one of embodiments 37-55, further comprisingadministering to the subject an inhibitor of IL-13, PDE4, or IL-31.

Embodiment 57

The method of embodiment 56, wherein if the subject has an altered geneexpression level of at least two of IL-13, IL-31, or TSLP, the subjectis administered with an inhibitor of IL-13, or the level of thetreatment is increased.

Embodiment 58

The method of embodiment 57, wherein the inhibitor of IL-13 islebrikizumab or tralokinumab.

Embodiment 59

The method of embodiment 56, wherein if the subject has an altered geneexpression level of at least two of IL-13, IL-31, or TSLP, the subjectis administered with an inhibitor of PDE4, or the level of the treatmentis increased.

Embodiment 60

The method of embodiment 56, wherein if the subject has an altered geneexpression level of at least two of IL-13, IL-3L or TSLP, the subject isadministered with an inhibitor of IL-31, or the level of the treatmentis increased.

Embodiment 61

A method of treating a subject with an antibody that specifically bindsto interleukin-13 (IL-13) or interleukin-13 receptor (IL-13R), whereinthe subject has atopic dermatitis, the method comprising the steps of:

-   -   determining whether the subject has an altered gene expression        level by:        -   obtaining or having obtained isolating nucleic acids from a            skin sample comprising cells from the stratum corneum; and        -   performing or having performed an expression analysis on the            skin sample by contacting the isolated nucleic acids with a            set of probes that recognizes at least two of IL-13, IL-31,            and TSLP, and detect binding between at least two of IL-13,            IL-31, and TSLP, and the set of probes; and    -   if the subject has an altered gene expression level of at least        two of IL-13, IL-31, and TSLP, then administer to the subject an        antibody that specifically binds to IL-13 or IL-13R, and    -   if the subject does not have an altered gene expression level of        at least two of IL-13, IL-31, and TSLP, then do not administer        the antibody that specifically binds to IL-13 or IL-13R.

Embodiment 62

The method of embodiment 61, wherein if the subject has an altered geneexpression of at least two of IL-13, IL-31, and TSLP, the subject isadministered an inhibitor of IL-13 or IL-13R, or the level of treatmentis increased.

Embodiment 63

The method of embodiment 62, wherein the altered gene expression is anincrease in expression.

Embodiment 64

The method of embodiment 61, wherein the antibody that specificallybinds to IL-13 is lebrikizumab or tralokinumab.

Embodiment 65

The method of embodiment 61, wherein the antibody that specificallybinds to IL-13R is dupilumab.

Embodiment 66

The method of any one of the embodiments 1-65, wherein the nucleic acidscomprise RNA, DNA, or a combination thereof.

Embodiment 67

The method of embodiment 66, wherein the RNA is mRNA.

Embodiment 68

The method of embodiment 66, wherein the RNA is cell-free circulatingRNA.

Embodiment 69

The method of any one of the embodiments 1-68, wherein the cells fromthe stratum corneum comprises T cells or components of T cells.

Embodiment 70

The method of any one of the embodiments 1-68, wherein the cells fromthe stratum corneum comprises keratinocytes.

Embodiment 71

The method of any one of the embodiments 1-70, wherein the skin sampleis obtained by applying an adhesive patch to a skin region of thesubject in a manner sufficient to adhere cells to the adhesive patch,and removing the adhesive patch from the skin region in a mannersufficient to retain the adhered cells to the adhesive patch.

Embodiment 72

The method of any one of the embodiments 1-70, wherein the skin sampleis obtained by applying a plurality of adhesive patches to a skin regionof the subject in a manner sufficient to adhere cells to each of theadhesive patches, and removing each of the adhesive patches from theskin region in a manner sufficient to retain the adhered cells to eachof the adhesive patches.

Embodiment 73

The method of embodiment 72, wherein the plurality of adhesive patchescomprises at least 4 adhesive patches.

Embodiment 74

The method of any one of the embodiments 1-73, wherein the amount ofnucleic acids isolated from the skin sample is from about 100 picogramsto about 100 micrograms, from about 200 picograms to about 10micrograms, or from about 500 picograms to about 1 microgram.

Embodiment 75

The method of any one of the embodiments 1-74, wherein the expressionlevel of genes is monitored over the course of 1 week, 2 weeks, 3 weeks,1 month, 2 months, 6 months, or more.

Embodiment 76

The method of any of the preceding embodiments, wherein the subject is ahuman.

While preferred embodiments of the present invention have been shown anddescribed herein, it will be obvious to those skilled in the art thatsuch embodiments are provided by way of example only. Numerousvariations, changes, and substitutions will now occur to those skilledin the art without departing from the invention. It should be understoodthat various alternatives to the embodiments of the invention describedherein may be employed in practicing the invention. It is intended thatthe following claims define the scope of the invention and that methodsand structures within the scope of these claims and their equivalents becovered thereby.

What is claimed is:
 1. A non-invasive method of detecting an autoimmunedisease and/or predicting flare-up, remission or a response to atreatment for, an autoimmune disease: a) collecting a tissue samplecomprising cellular material from a non-lesional area of skin from asubject suspected of having the autoimmune disease using an adhesivepatch sampling of the cellular material from the stratum corneum onto anadhesive patch, wherein the cellular material comprises nucleic acids,b) isolating nucleic acids from the tissue sample collected from thesubject, and c) detecting the autoimmune disease and/or predictingflare-up, remission or response to a treatment for, the autoimmunedisease based on expression levels derived from the isolated nucleicacids.
 2. The method of claim 1, wherein (i) the adhesive patch samplingremovably adheres the cellular material from the stratum corneum onto anadhesive material on the adhesive patch and/or (ii) the type of cellularmaterial obtained from the adhesive patch sampling comprises cellsobtained from the skin no deeper than the stratum corneum.
 3. The methodof claim 1, wherein an amount of cellular material collected by theadhesive patch sampling is no more than about 1 gram.
 4. The method ofclaim 1, wherein the expression levels of nucleic acids are derived fromone or more genes in a gene classifier associated with the immune and/orinflammatory pathways elevated in atopic dermatitis, lupus, orpsoriasis.
 5. The method of claim 4, wherein the pathways comprise Th1,Th2, Th17, or Th22.
 6. The method of claim 1, wherein expression levelsof one or more genes from a gene classifier associated with atopicdermatitis are analyzed from the sample, and the gene classifiercomprises: (i) IL-13, IL-31, or TSLP; (ii) IL-13R, IL-4R, IL-17, IL-22,CXCL9, CXCL10, CXCLH, S100A7, S100A8, S100A9, CCL17, CCL18, CCL19,CCL26, CCL27, or NOS2; (iii) IL-31RA, CCL17, IL-23A, IL-4R, IL22, IL-13,or IL-13RA1; or (iv) IL-13 pathway constituents or receptors.
 7. Themethod of claim 1, comprising predicting a response to a treatment forthe autoimmune disease, wherein the autoimmune disease comprises atopicdermatitis, and the treatment comprises an inhibitor of IL-13, aninhibitor of PDE4, or an inhibitor of IL-31.
 8. The method of claim 1,comprising predicting a response to a treatment for the autoimmunedisease, wherein the autoimmune disease comprises atopic dermatitis, andthe treatment comprises lebrikizumab, tralokinumab, crisaborole, ornemolizumab.
 9. The method of claim 1, wherein expression levels of oneor more genes from a gene classifier associated with psoriasis areanalyzed from the sample, and the gene classifier comprises: (i) IL-17A,IL-17F, IL-8, CXCL5, S100A9, or DEFB4A; (ii) IL-17C, S100A7, IL-17RA,IL-17RC, IL-23A, IL-22, IL-26, IL-24, IL-6, CXCL1, IFN-gamma, IL-31,IL-33, TNFa, LCN2, CCL20, or TNFRSF1A; or (iii) IL-17A, IL-17C, IL-17F,IL-17 receptor, IL-23 A, IL-22, IL-24, IL-6, IL-8, CXCL1, CXCL5, DEFB4A,LCN2, S100A7, TNF-alpha, or TNF-alpha receptor.
 10. The method of claim1, comprising predicting a response to a treatment for the autoimmunedisease, wherein the autoimmune disease comprises psoriasis, and thetreatment comprises an inhibitor of TNF-alpha, an inhibitor of IL-17A,or an inhibitor of IL-23.
 11. The method of claim 1, comprisingpredicting a response to a treatment for the autoimmune disease, whereinthe autoimmune disease comprises psoriasis, and the treatment comprisesadalimumab, certolizumab, etanercept, golimumab, and infliximab,ixekizumab (LY2439821), brodalumab (AMG 827), secukinumab, guselkumab,tildrakizumab, or risankizumab.
 12. The method of claim 1, whereinexpression levels of one or more genes from a gene classifier associatedwith lupus are analyzed from the sample, and the gene classifiercomprises: (i) IFNAJ, IFNA2, IFNA4, 11 ′NR 1, IFNR2, CCL5; or (ii)IFNB1, IFNE, IFNWI, ADAR, IFIT, IFI, IRF, OAS1, TRAM, TNFAIP3, ATG5,TYK2, STAT4, OPN, or KRT.
 13. The method of claim 1, comprisingpredicting a response to a treatment for the autoimmune disease, whereinthe autoimmune disease comprises lupus, and the treatment comprises: anantimalarial, dapsone, a retinoid, a corticosteroid, animmunosuppressive drug, thalidomide, a Janus kinase inhibitor, Dapsone,baricitinib, hydroxychloroquine, quinacrine, chloroquine, methotrexate,or azathioprine.
 14. The method of claim 1, wherein detecting thepresence of, and/or predicting a response to a treatment for, theautoimmune disease comprises: contacting the isolated nucleic acids witha set of probes that recognize one or more genes involved in thecytokine-mediated immune and inflammatory responses, detecting ormeasuring an amount of binding between the nucleic acids and the set ofprobes, and comparing the amount of binding between the nucleic acidsand the set of probes in the sample relative to a control or thresholdamount of binding.
 15. The method of claim 14, comprising administeringto the subject the treatment for the autoimmune disease when the amountof binding between the nucleic acids and the set of probes is altered inthe sample relative to the control or threshold amount of binding.
 16. Amethod for non-invasively identifying an autoimmune disease, comprising:a) identifying a subject suspected of having an autoimmune disease, thesubject having lesional and non-lesional areas of skin; b) applying anadhesive patch to a non-lesional area of the subject's skin in a mannersufficient to adhere a sample of cellular material from the stratumcorneum to the adhesive patch, wherein the sample of cellular materialcomprises nucleic acids; c) removing the adhesive patch from thesubject's skin in a manner sufficient to retain the sample of cellularmaterial adhered to the adhesive patch; and d) detecting an autoimmunedisease and/or predicting flare-up, remission or a response to atreatment for, the autoimmune disease based on expression levels derivedfrom the isolated nucleic acids.
 17. The method of claim 16, wherein theautoimmune disease comprises atopic dermatitis, and expression levels ofone or more genes from a gene classifier are analyzed from the sample,wherein the gene classifier comprises: (i) IL-13, IL-31, or TSLP; (ii)IL-13R, IL-4R, IL-17, IL-22, CXCL9, CXCL10, CXCLH, S100A7, S100A8,S100A9, CCL17, CCL18, CCL19, CCL26, CCL27, or NOS2; (iii) IL-31RA,CCL17, IL-23A, IL-4R, IL22, IL-13, or IL-13RA1; or (iv) IL-13 pathwayconstituents or receptors.
 18. The method of claim 16, wherein theautoimmune disease comprises psoriasis, and expression levels of one ormore genes from a gene classifier are analyzed from the sample, whereinthe gene classifier comprises: (i) IL-17A, IL-17F, IL-8, CXCL5, S100A9,or DEFB4A; (ii) IL-17C, S100A7, IL-17RA, IL-17RC, IL-23A, IL-22, IL-26,IL-24, IL-6, CXCL1, IFN-gamma, IL-31, IL-33, TNFa, LCN2, CCL20, orTNFRSF1A; or (iii) IL-17A, IL-17C, IL-17F, IL-17 receptor, IL-23 A,IL-22, IL-24, IL-6, IL-8, CXCL1, CXCL5, DEFB4A, LCN2, S100A7, TNF-alpha,or TNF-alpha receptor.
 19. The method of claim 16, wherein theautoimmune disease comprises lupus, and expression levels of one or moregenes from a gene classifier are analyzed from the sample, wherein thegene classifier comprises: (i) IFNAJ, IFNA2, IFNA4, 11 ′NR 1, IFNR2,CCL5; or (ii) IFNB1, IFNE, IFNWI, ADAR, IFIT, IFI, IRF, OAS1, TRAM,TNFAIP3, ATG5, TYK2, STAT4, OPN, or KRT.
 20. The method of claim 16,wherein a) the autoimmune disease comprises atopic dermatitis, and thetreatment comprises: an inhibitor of IL-13, an inhibitor of PDE4, aninhibitor of IL-31, lebrikizumab, tralokinumab, crisaborole, ornemolizumab; b) the autoimmune disease comprises psoriasis, and thetreatment comprises: an inhibitor of TNF-alpha, an inhibitor of IL-17A,an inhibitor of IL-23, adalimumab, certolizumab, etanercept, golimumab,and infliximab, ixekizumab (LY2439821), brodalumab (AMG 827),secukinumab, guselkumab, tildrakizumab, or risankizumab; or c) theautoimmune disease comprises lupus, and the treatment comprises: anantimalarial, dapsone, a retinoid, a corticosteroid, animmunosuppressive drug, thalidomide, a Janus kinase inhibitor, Dapsone,baricitinib, hydroxychloroquine, quinacrine, chloroquine, methotrexate,or azathioprine.